hTERT Gene Research Articles

Adipose Tissue-Mesenchymal Stem Cells Caused to Change the Methylation Status of hTERT Gene Promoter CpG Islands of Molt-4 Leukemia Cells as Cell-based Therapy.

DNA methylation was considered as prognostic information in some hematological malignancies. Previous studies have reported the in vitro and in vivo biology role of mesenchymal stem cells (MSCs) on leukemic cells. The aim of this study was to investigate the effect of MSCs on the promoter methylation status of hTERT as a catalytic subunit of telomerase enzyme. In the experimental study, the Molt-4 leukemic cells were co-cultured with MSCs for 7 days. At the end of the co-culture period, the Molt-4 cells were collected, DNA and protein were extracted. Then methylation specific-PCR and western blotting were done for evaluating the hTERT gene promoter methylation status and cyclin D1 and hTERT protein expression, respectively. In the following, the flow cytometry was done for cell cycle distribution assay. It was found that MSCs resulted in a significant decrease in the cyclin D1 and hTERT protein expression levels. Also, MSCs caused changes in the methylation status of the CpG islands in the hTERT gene promoter region. The following results showed that MSCs caused a significant increase in the number of cells at G0/G1 phase and arrest the G0/G1 phase as well as decrease in the cell proliferation of Molt-4 cells. It is concluded that co-culture of MSCs with Molt-4 cells could be involved in changing the methylation status of hTERT gene promoter, cell cycle and hTERT protein expression; it could be potentially beneficial for further investigations regarding the cell transplantation and cell-based therapy.

Telomere Length as a New Risk Marker of Early-Onset Colorectal Cancer.

Early-onset colorectal cancer (EOCRC; age younger than 50 years) incidence has been steadily increasing in recent decades worldwide. The need for new biomarkers for EOCRC prevention strategies is undeniable. In this study, we aimed to explore whether an aging factor, such as telomere length (TL), could be a useful tool in EOCRC screening. The absolute leukocyte TL from 87 microsatellite stable EOCRC patients and 109 healthy controls (HC) with the same range of age, was quantified by Real Time Quantitative PCR (RT-qPCR). Then, leukocyte whole-exome sequencing (WES) was performed to study the status of the genes involved in TL maintenance (hTERT, TERC, DKC1, TERF1, TERF2, TERF2IP, TINF2, ACD, and POT1) in 70 sporadic EOCRC cases from the original cohort. We observed that TL was significantly shorter in EOCRC patients than in healthy individuals (EOCRC mean: 122 kb vs. HC mean: 296 kb; p < 0.001), suggesting that telomeric shortening could be associated with EOCRC susceptibility. In addition, we found a significant association between several SNPs of hTERT (rs79662648), POT1 (rs76436625, rs10263573, rs3815221, rs7794637, rs7784168, rs4383910, and rs7782354), TERF2 (rs251796 and rs344152214), and TERF2IP (rs7205764) genes and the risk of developing EOCRC. We consider that the measurement of germline TL and the status analysis of telomere maintenance related genes polymorphisms at early ages could be non-invasive methods that could facilitate the early identification of individuals at risk of developing EOCRC.

Spotlight on hTERT Complex Regulation in Cutaneous T-Cell Lymphomas

As a major cancer hallmark, there is a sustained interest in understanding the telomerase contribution to carcinogenesis in order to therapeutically target this enzyme. This is particularly relevant in primary cutaneous T-cell lymphomas (CTCL), a malignancy showing telomerase dysregulation with few investigative data available. In CTCL, we examined the mechanisms involved in telomerase transcriptional activation and activity regulation. We analyzed 94 CTCL patients from a Franco-Portuguese cohort, as well as 8 cell lines, in comparison to 101 healthy controls. Our results showed that not only polymorphisms (SNPs) located at the promoter of human telomerase reverse transcriptase (hTERT) gene (rs2735940 and rs2853672) but also an SNP located within the coding region (rs2853676) could influence CTCL occurrence. Furthermore, our results sustained that the post-transcriptional regulation of hTERT contributes to CTCL lymphomagenesis. Indeed, CTCL cells present a different pattern of hTERT spliced transcripts distribution from the controls, mostly marked by an increase in the hTERT β+ variants proportion. This increase seems to be associated with CTCL development and progression. Through hTERT splicing transcriptome modulation with shRNAs, we observed that the decrease in the α-β+ transcript induced a decrease in the cell proliferation and tumorigenic capacities of T-MF cells in vitro. Taken together, our data highlight the major role of post-transcriptional mechanisms regulating telomerase non canonical functions in CTCL and suggest a new potential role for the α-β+ hTERT transcript variant.

Potential Anti-Proliferative Effect of Nano-formulated Curcumin Through Modulating Micro RNA- 132, Cyclin D1, and hTERT Genes Expression in Breast Cancer Cell Lines

Potential Anti-Proliferative Effect of Nano-formulated Curcumin Through Modulating Micro RNA- 132, Cyclin D1, and hTERT Genes Expression in Breast Cancer Cell Lines

Establishment and characterization of an immortalized bovine intestinal epithelial cell line.

Primary bovine intestinal epithelial cells (PBIECs) are an important model for studying the molecular and pathogenic mechanisms of diseases affecting the bovine intestine. It is difficult to obtain and grow PBIECs stably, and their short lifespan greatly limits their application. Therefore, the purpose of this study was to create a cell line for exploring the mechanisms of pathogen infection in bovine intestinal epithelial cells in vitro. We isolated and cultured PBIECs and established an immortalized BIEC line by transfecting PBIECs with the pCI-neo-hTERT (human telomerase reverse transcriptase) recombinant plasmid. The immortalized cell line (BIECs-21) retained structure and function similar to that of the PBIECs. The marker proteins characteristic of epithelial cells, cytokeratin 18 (CK18), occludin, zonula occludens protein 1 (ZO-1), E-cadherin and enterokinase, were all positive in the immortalized cell line, and the cell structure, growth rate, karyotype, serum dependence and contact inhibition were normal. The hTERT gene was successfully transferred into BIECs-21 where it remained stable and was highly expressed. The transport of short-chain fatty acids and glucose uptake by the BIECs-21 was consistent with PBIECs, and we showed that they could be infected with the intestinal parasite, Neospora caninum. The immortalized BIECs-21, which have exceeded 80 passages, were structurally and functionally similar to the primary BIECs and thus provide a valuable research tool for investigating the mechanism of pathogen infection of the bovine intestinal epithelium in vitro.

Establishing an hTERT-driven immortalized umbilical cord-derived mesenchymal stem cell line and its therapeutic application in mice with liver failure

Acute liver failure (ALF) is characterized by rapid liver cell destruction. It is a multi-etiological and fulminant complication with a clinical mortality of over 80%. Therapy using mesenchymal stem cells (MSCs) or MSCs-derived exosomes can alleviate acute liver injury, which has been demonstrated in animal experiments and clinical application. However, similar to other stem cells, different cell sources, poor stability, cell senescence and other factors limit the clinical application of MSCs. To achieve mass production and quality control on stem cells and their exosomes, transfecting umbilical cord mesenchymal stem cell (UCMSC) with lentivirus overexpressing human telomerase reverse transcriptase (hTERT) gene, the hTERT-UCMSC was constructed as an immortalized MSC cell line. Compared with the primary UCMSC (P3) and immortalized cell line hTERT-UCMSC at early passage (P10), the hTERT-UCMSC retained the key morphological and physiological characteristics of UCMSC at the 35th passage (P35), and showed no signs of carcinogenicity and toxic effect in mice. There was no difference in either exosome production or characteristics of exosomes among cultures from P3 primary cells, P10 and P35 immortalized hTERT-UCMSCs. Inoculation of either hTERT-UCMSC (P35) or its exosomes improved the survival rate and liver function of ALF mice induced by thioacetamide (TAA). Our findings suggest that this immortalized cell line can maintain its characteristics in long-term culture. Inoculation of hTERT-UCMSC and its exosomes could potentially be used in clinics for the treatment of liver failure in the future.

Potential Anti-Cancer Effect of Helenalin as a Natural Bioactive Compound on the Growth and Telomerase Gene Expression in Breast Cancer Cell Line.

The telomerase gene is overexpressed in the majority of tumors and cancers compared to normal and healthy cells, and on the other hand, this enzymatic protein is overactive, therefore, the telomerase enzyme is considered a primary target for diagnostic and therapeutic purposes in most cancers. This has been hypothesized that Helenalin has anti-telomerase activity in a wide range of cancers and Tumor tissues. In this study, we investigated the inhibitory effect of helenalin extract on telomerase gene expression in the T47D breast cancer cell line. We used the MTT assay to evaluate the cytotoxic effect of different concentrations of helenalin on the T47D breast cancer cell line at 24, 48, and 72 hours. Besides, the expression of the hTERT gene in T47D cell lines treated with 1.0 and 5.0 µM helenalin after 24, 48, and 72 h incubation times was investigated through real-time PCR. According to the MTT assay, the inhibitory effect of helenalin on T47D cell proliferation is time and dose-dependent. Moreover, the results of Real-time PCR showed that exposure of T47D cell lines to helenalin led to a significant Decreasing in the expressional values of the hTERT gene as a time and dose-dependent procedure compared with the control group (P ≤ 0.05). These preliminary results demonstrated the cytotoxic potential of helenalin through inhibition of hTERT against T47D breast cancer cells.

Telomerase inhibition in malignant gliomas. A systematic review.

Glioblastoma (GBM) is the most frequent adult malignant brain tumour and despite different therapeutic efforts, the median overall survival still ranges from 14 to 18 months. Thus, new therapeutic strategies are urgently needed. However, the identification of cancer-specific targets is particularly challenging in GBM, due to the high heterogeneity of this tumour in terms of histopathological, molecular, genetic and epigenetic features. Telomerase reactivation is a hallmark of malignant glioma. An activating mutation of the hTERT gene, encoding for the active subunit of telomerase, is one of the molecular criteria to establish a diagnosis of GBM, IDH-wildtype, in the 2021 WHO classification of central nervous system tumours. Telomerase inhibition therefore represents, at least theoretically, a promising strategy for GBM therapy: pharmacological compounds, as well as direct gene expression modulation therapies, have been successfully employed in in vitro and in vivo settings. Unfortunately, the clinical applications of telomerase inhibition in GBM are currently scarce. The aim of the present systematic review is to provide an up-to-date report on the studies investigating telomerase inhibition as a therapeutic strategy for malignant glioma in order to foster the future translational and clinical research on this topic.

A combination of telomerase inhibition and NK cell therapy increased breast cancer cell line apoptosis

Triple-negative breast cancer (TNBC) is a subtype of breast tumor with the highest breast cancer stem cells (BCSCs) content and resistance to conventional treatment. Due to the immunosuppressive tumor microenvironment and immunogenicity of breast cancer cells, the use of immune cells, especially natural killer cells (NK) in the treatment of solid tumors, including breast cancer, has been unsatisfactory. Therefore, identifying novel therapies is requisite for breast cancer treatment. Furthermore, the combination of cancer therapies is an effective strategy to improve therapeutic effectiveness. In this study, we inhibited telomerase (hTERT) with BIBR1532, in stimulating NK cell cytotoxicity against breast cancer cells. The MDA-MB-231 cell line was cured with IC50 level of BIBR1532 for 24 h. Afterward, cells were washed with PBS and were co-cultured with peripheral blood NK cell for 5h. Finally, we assessed the impact of telomerase inhibition on the cytotoxicity of NK cells and apoptosis of breast cancer. Also, the expression of hTERT and apoptotic-related genes were evaluated. The data revealed that inhibition of telomerase increases NK cell cytotoxicity against breast cancer. Furthermore, telomerase inhibition and NK cell synergistically enhanced cell death in breast cancer cells by suppressing hTERT, upregulation of bax, and bad expression. In conclusion, telomerase suppression makes breast cancer cells more sensitive to NK cell therapy. Consequently, the combination of telomerase inhibition and NK cells can be useful in the treatment of breast cancer cells.

Midazolam alleviates cellular senescence in SH-SY5Y neuronal cells in Alzheimer's disease.

Alzheimer's disease (AD) impacts the daily life of aging people. Oligomerized amyloid β (Aβ)-associated neuronal senescence is involved in the pathological mechanism of AD. Blockage of neuronal senescence has been considered an important strategy for the treatment of AD. Midazolam is a hypnotic-sedative drug with pleiotropic properties. However, the effects of Midazolam in oligomerized Aβ1.42 -induced neurotoxicity have not been reported previously. Here, we investigate whether Midazolam possesses a beneficial effect against oligomerized Aβ1.42 in SH-SY5Y neuronal cells. Cellular senescence was assessed using senescence-associated β-galactosidase staining. Telomerase activity was measured using the TeloTAGGG Telomerase PCR ELISA. First, the lactate dehydrogenase release assay demonstrates that 10 and 20 µM are the optimal concentrations of Midazolam used for cell cultures. Senescence-associated β-galactosidase staining results indicate that exposure to oligomerized Aβ1.42 significantly increased cellular senescence of SH-SY5Y cells, but it was significantly alleviated by Midazolam. Additionally, Midazolam restored the oligomerized Aβ1.42 -induced reduction of telomerase activity. Interestingly, we found that oligomerized Aβ1.42 remarkably reduced human telomerase reverse transcriptase (hTERT) gene expression but increased the telomeric repeat-binding factor 2 (TERF2) expression. However, treatment with Midazolam reversed the effects of oligomerized Aβ1.42 on the hTERT and TERF2 gene expressions. Importantly, the presence of Midazolam attenuated Aβ1.42 -induced p53 and p21 expressions. Mechanistically, Midazolam repressed the level of cyclooxygenase-2 (COX-2) and the release of prostaglandin E2. Importantly, overexpression of COX-2 abolished the impact of Midazolam against oligomerized Aβ1.42 in neuronal senescence. We conclude that the usage of Midazolam is a potential treatment strategy for AD.

Implications of risk conferred by 5p15.33 loci genetic variants; human telomerase reverse transcriptase rs2736098 and rs2736100 in predisposition of bladder cancer.

The polymorphic variations of human telomerase reverse transcriptase (hTERT) gene play an important role in predisposition to carcinogenesis. The current study aimed to elucidate the genetic predisposition to bladder cancer in two important variants, rs2736098 and rs2736100 of hTERT gene. Confirmed 130 patients of bladder cancer and 200 healthy controls were genotyped by PCR-RFLP to determine different variants of hTERT rs2736098 and rs2736100. hTERT rs2736098 homozygous variant AA genotype frequency was observed to significantly differ 2-fold between cases and controls (26.15% vs. 13.5%) (p = 0.02). In addition, rare 'A' allele significantly differed among two groups (cases: 47% versus controls: 39%: p = 0.03). hTERT rs2736098 was observed to be presented significantly more in high stage tumors (p = 0.02). hTERT rs2736100 genotype AA or variant allele A showed no significant difference between cases and controls. Haplotype CA displayed significantly different pattern of frequency as 0.5 in cases as compared to 0.16 in controls (p < 0.0001). Combination of variant A/G haplotype frequency implicated more in cases than in controls (0.34 vs. 0.14, p = 0.001). It is concluded that hTERT rs2736098 polymorphic variant has a vital role to confer a strong risk to bladder cancer in our population. Further, hTERT haplotypes CA and AG inhTERT could prove to be a promising tool to screen the risk for bladder cancer.

The effect of natural polyphenols Resveratrol, Gallic acid, and Kuromanin chloride on human telomerase reverse transcriptase (hTERT) expression in HepG2 hepatocellular carcinoma: role of SIRT1/Nrf2 signaling pathway and oxidative stress.

There is evidence that low doses or physiological concentrations of certain natural polyphenols enhance the activity of telomerase. However, the precise mechanism by which natural polyphenols regulate telomerase activity remains unclear. Recent research indicates that NF-E2 related factor 2 (Nrf2) and silent information regulator 1 (SIRT1) are involved in human telomerase reverse transcriptase (hTERT) regulation. Thus, in order to better comprehend the mechanism by which polyphenols regulate hTERT, the present study investigated the effects of the natural polyphenols Resveratrol, Gallic acid, and Kuromanin chloride on hTERT, Nrf2, and SIRT1 expression as well as oxidative stress in HepG2 hepatocellular carcinoma. The trypan blue dye exclusion assay was used to assess cell viability. The level of mRNA for hTERT, Nrf2, and SIRT1 was then determined using real-time PCR. A spectrophotometric analysis was conducted to quantify oxidative stress markers. The results demonstrated that Resveratrol induces the expression of hTERT and the SIRT1/Nrf2 pathway in a dose-dependent manner. Gallic acid at concentrations of 10 and 20μM also increased the expression of the hTERT and SIRT1/Nrf2 pathway. Furthermore, dose-dependent overexpression of hTERT and Nrf2 was induced by Kuromanin chloride at 10 and 20µM. Moreover, we found that Resveratrol and Kuromanin chloride ameliorated oxidative stress, whereas Gallic acid exacerbated it. This study demonstrates that low doses of polyphenols (Resveratrol, Gallic acid, and Kuromanin chloride) upregulate the expression of the hTERT gene in the HepG2 hepatocellular carcinoma cell line, possibly via induction of the SIRT1/Nrf2 signaling pathway. Therefore, by targeting this pathway or hTERT, the anti-cancer effect of polyphenols can be enhanced.

Antiproliferative Effects of Ferulic, Coumaric, and Caffeic Acids in HepG2 Cells by hTERT Downregulation.

Phenolic acids are well-known phytochemicals that are detected in a wide variety of medicinal plants, and their antiproliferative effects on cancer cells are known, but their mechanisms are poorly revealed. In most of cancer cells, telomerase reverse transcriptase (hTERT) is a dominant factor of telomere length regulation. The hTERT expression promotes invasiveness in tumor cells and is a hallmark of cancer. Therefore, in this study, the probable inhibitory effects of caffeic (Caf), coumaric (Cum), and ferulic acids (Fer) are investigated on the hTERT expression pattern in HepG2 cells. The MTT, apoptosis assays, and real-time PCR analysis were applied to evaluate viability, cytotoxicity, and hTERT gene expression level, respectively. All of the studied phenolic acids showed cytotoxic effects on HepG2 cells in a timely manner and presented a time-dependent inhibitory effect on the growth of HepG2 cells. They reduced percentage of viable cells and induced apoptosis. Also, these phenolic acids had significant inhibitory effects on hTERT gene expression. These findings suggest that cell viability along with hTERT gene expression in HepG2 cells could be reduced by Cum, Caf, and Fer. As different cancer cells are resistant to conventional chemotherapeutics, this type of results proposes the telomerase as a proper target of cancer therapy development by natural products.

Anticancer activity, hTERT expression and telomere length analysis in SH-SY5Y neuroblastoma cell lines applied to docetaxel

Neuroblastoma is the most common extracranial solid tumor of infancy in a broad range of clinical courses, ranging from spontaneous regression to fatal progression. Telomere maintenance plays an important role in genome stability and cell proliferation. Telomerase reverse transcriptase (hTERT in humans) is a catalytic subunit of the enzyme telomerase. In this study, it was aimed to determine the anticancer activity of the docetaxel chemotherapeutic agent in neuroblastoma cell line (SH-SY5Y) and to investigate its effect on hTERT gene expression level and telomere length. Molecular docking studies were performed on docetaxel with the crystal structure of telomerase. The electronic properties of docetaxel were calculated using the density functional theory (DFT) method. SH-SY5Y cells were treated for 24, 48 and 72 h with specific concentrations of docetaxel drug ranging from 1 to 100 µg/ml. IC50 doses of docetaxel were determined and administered to SH-SY5Y cells, followed by RNA isolation. hTERT and MYC gene expression levels and telomere length were measured in the docetaxel-treated sample using the RT-PCR method. In addition, theoretical analyzes were made. The IC50 values of docetaxel after 24, 48 and 72 h were 8.32±1.45 μg/ml, 7.67±2.56 μg/ml and 5.51±1.24 μg/ml, respectively. According to the results obtained, docetaxel was found to have the highest activity in 72 h of incubation. It was determined that the docetaxel drug decreased the expression level of the hTERT gene in SH-SY5Y cells. Telomere lengths were significantly reduced in the docetaxel treated SH-SY5Y cell line compared to the control group (p < 0.05). Molecular docking analysis results were in agreement with the experiments. Analysis results indicated a good interaction between docetaxel and the active site of telomerase. The results of this study, reinforced by molecular docking analyzes, might be proved valuable for the development of potent telomerase inhibitors.

Telomerase Reverse Transcriptase and Telomerase RNA Component Gene Expression as a Novel Biomarkers for Alzheimer's disease.

Alzheimer's disease (AD) is a neurological, age-related condition that causes cognitive decline and memory loss; it induces dementia in the elderly. Telomerase is a reverse transcriptase ribonucleoprotein that adds nucleotides to the end of DNA. This study aimed to compare human telomerase reverse transcriptase (hTERT) and telomerase RNA component (TERC) expression in different phases of AD and healthy cohorts. Sixty participants were divided into 30 who had dementia and 30 who did not. After collecting blood samples, total RNAs were extracted from the plasma. Screening for hTERT and TERC gene expression was carried out by quantitative reverse transcriptase real-time polymerase chain reaction (RT-qPCR) using the relative quantification method to estimate the expression changes in hTERT and TERC. The RT-qPCR results show that hTERT and TERC gene expression was significantly down-regulated in Alzheimer's patients compared to the health subjects (P-value= <0.0001,0.005), respectively. The area under curve AUC was 0.773 for hTERT and 0.703 for TERC. The Mini-Mental State Examination scores revealed a significant difference between dementia and non-dementia subjects (P=<0.0001). We conclude down-regulations in both hTERT and TERC gene expression in AD patients, which supports our hypothesis that the telomerase expression gene in the blood of AD patients can serve as a non-invasive, early, and novel diagnostic marker of AD.

AST-487 Inhibits RET Kinase Driven TERT Expression in Bladder Cancer.

Mutations in the promoter of the human Telomerase Reverse Transcriptase (hTERT) gene are common and associated with its elevated expression in bladder cancer, melanoma, and glioblastoma. Though these mutations and TERT overexpression are associated with aggressive disease and poor outcome, an incomplete understanding of mutant TERT regulation limits treatment options directed at this gene. Herein, we unravel a signaling pathway that leads to upregulated hTERT expression resulting from the −124 bp promoter mutation, the most frequent variant across human cancer. We employed engineered bladder cancer cells that harbor a GFP insertion at the TSS region on −124 hTERT promoter for high-content screening drug discovery using a focused library of ~800 kinase inhibitors. Studies using in vitro and in vivo models prioritized AST-487, an inhibitor of the wild-type, and mutant RET (rearranged during transfection) proto-oncogene as a novel drug inhibitor of both wild-type and mutant promoter-driven hTERT expression. We also identified the RET kinase pathway, targeted by AST-487, as a novel regulator of mutant hTERT promoter-driven transcription in bladder cancer cells. Collectively, our work provides new potential precision medicine approaches for cancer patients with upregulated hTERT expression, perhaps, especially those harboring mutations in both the RET gene and the hTERT promoter, such as in thyroid cancer.

Differential DNA Methylation of THOR and hTAPAS in the Regulation of hTERT and the Diagnosis of Cancer.

Simple SummaryBecause of its high prevalence of >45% in 9 out of 11 (82%) cancer types screened, THOR hypermethylation has been suggested to be a frequent telomerase-activating mechanism in hTERT-expressing tumor types, e.g., in cancers of the prostate, breast, blood, colon, lung, bladder, and brain. In this prime example, we present detailed DNA methylation profiles in urothelial cancer that reveal the exact positions of the most differentially methylated CpG dinucleotides within the THOR region in order to design an efficient Methylation-Specific PCR (MSPCR) approach for diagnostic and prognostic purposes. Furthermore, our data suggest an epigenetic mechanism regulating hTERT expression through the methylation status of THOR and lncRNA hTAPAS.Background: Although DNA methylation in the gene promoters usually represses gene expression, the TERT hypermethylated oncological region (THOR) located 5′ of the hTERT gene is hypermethylated when hTERT is expressed in diverse cancer types, including urothelial cancer (UC). Methods: Comprehensive MeDIP and DNA methylation array analyses complemented by the technically independent method of bisulfite genomic sequencing were applied on pathologically reviewed and classified urothelial carcinoma specimens and healthy urothelial tissue samples to reveal the methylation status of THOR in detail. Results: The detailed DNA methylation profiles reveal the exact positions of differentially methylated CpG dinucleotides within THOR in urothelial cancer and provide evidence ofa diverging role of methylation of these CpGs in the regulation of hTERT. In particular, our data suggest a regulating mechanism in which THOR methylation acts on hTERT expression through epigenetic silencing of the lncRNA hTERT antisense promoter-associated (hTAPAS), which represses hTERT. Conclusions: These findings precisely define the most differentially methylated CpGs of THOR in early urothelial cancer, enabling optimal design of Methylation-Specific PCR (MSPCR) primers to reliably probe these methylation differences for diagnostic and prognostic purposes. In addition, this strategy presents a prime example that is also applicable to many other malignancies. Finally, the first evidence for the underlying epigenetic mechanism regulating hTERT expression through the methylation status of THOR is provided.

Antisenescence Effect of REAC Biomodulation to Counteract the Evolution of Myelodysplastic Syndrome

About 30 percent of patients diagnosed with myelodysplastic syndromes (MDS) progress to acute myeloid leukemia (AML). The senescence of bone marrow?derived mesenchymal stem cells (BMSCs) seems to be one of the determining factors in inducing this drift. Research is continuously looking for new methodologies and technologies that can use bioelectric signals to act on senescence and cell differentiation towards the phenotype of interest. The Radio Electric Asymmetric Conveyer (REAC) technology, aimed at reorganizing the endogenous bioelectric activity, has already shown to be able to determine direct cell reprogramming effects and counteract the senescence mechanisms in stem cells. Aim of the present study was to prove if the anti-senescence results previously obtained in different kind of stem cells with the REAC Tissue optimization - regenerative (TO-RGN) treatment, could also be observed in BMSCs, evaluating cell viability, telomerase activity, p19ARF, P21, P53, and hTERT gene expression. The results show that the REAC TO-RGN treatment may be a useful tool to counteract the BMSCs senescence which can be the basis of AML drift. Nevertheless, further clinical studies on humans are needed to confirm this hypothesis.

Establishment of a fetal cow (Bos Borus) skin fibroblasts cell line with immortalized characterization through human telomerase reverse transcriptase (hTERT) ectopic expression

The ectopic introduction of the human telomerase reverse transcriptase (hTERT) is an effective way to establish an immortalized cell line. Here, hTERT was obtained by RT-PCR, and the eukaryotic expression plasmid and lentivirus shuttle plasmid of hTERT was successfully constructed by the homologous recombination method. The stable expression of hTERT in fetal cow skin fibroblasts (CSF) was established using the lentivirus package system. The hTERT-CSF proliferate and have immortalized characteristics. Meanwhile, the chromosome analysis identified that the number and structure of the hTERT-CSF genome maintain stable. The indirect immunofluorescence, western blot, and flow cytometry showed that the hTERT gene had been successfully integrated into the primary genome of bovine skin and stably expressed. The viral infection experiment first identifies the hTERT-CSF as a vulnerable cell model responding to the Lumpy skin disease virus (LSDV). Establishing hTERT-CSF provides an important cell model for basic and applied research, clinical application, and vaccine development. It provides an essential reference for the future’s rapid establishment of other immortalized cell lines.

Panic disorder aging characteristics: The role of telomerase reverse transcriptase gene and brain function.

Panic disorder (PD) causes serious functional damage and disability and accelerates the process of individual aging. The pathological basis of PD is the same as that of age-related diseases, which is proposed as a new viewpoint in recent years. Memory decline and social functional impairment are common manifestations of accelerated aging in PD. The function of telomerase reverse transcriptase (TERT) and telomere length (TL) is abnormal in patients with aging and PD. However, the molecular mechanism behind remains unclear. The purpose of this study was to explore the relationship between TERT gene expression (including DNA methylation) and the changes in PD aging characteristics (memory and social function). By TERT gene knockout mice, we found that loss of TERT attenuated the acquisition of recent fear memory during contextual fear conditioning. This study reported that a significantly lower methylation level of human TERT (hTERT) gene was detected in PD patients compared with healthy control and particularly decreased CpG methylation in the promoter region of hTERT was associated with the clinical characteristics in PD. Regional homogeneity (ReHo) analysis showed that the methylation of hTERT (cg1295648) influenced social function of PD patients through moderating the function of the left postcentral gyrus (PCG). This indicates that the hTERT gene may play an important role in the pathological basis of PD aging and may become a biological marker for evaluating PD aging. These findings provide multidimensional evidence for the underlying genetic and pathological mechanisms of PD.