Telomerase-related advances in hepatocellular carcinoma:A bibliometric and visual analysis

BACKGROUND As a critical early event in hepatocellular carcinogenesis,telomerase activation might be a promising and critical biomarker for hepatocellular carcinoma(HCC)patients,and its function in the genesis and treatment of HCC has gained much attention over the past two decades.AIM To perform a bibliometric analysis to systematically assess the current state of research on HCC-related telomerase.METHODS The Web of Science Core Collection and PubMed were systematically searched to retrieve publications pertaining to HCC/telomerase limited to“articles”and“reviews”published in English.A total of 873 relevant publications related to HCC and telomerase were identified.We employed the Bibliometrix package in R to extract and analyze the fundamental information of the publications,such as the trends in the publications,citation counts,most prolific or influential writers,and most popular journals;to screen for keywords occurring at high frequency;and to draw collaboration and cluster analysis charts on the basis of coauthorship and co-occurrences.VOSviewer was utilized to compile and visualize the bibliometric data.RESULTS A surge of 51 publications on HCC/telomerase research occurred in 2016,the most productive year from 1996 to 2023,accompanied by the peak citation count recorded in 2016.Up to December 2023,35226 citations were made to all publications,an average of 46.6 citations to each paper.The United States received the most citations(n=13531),followed by China(n=7427)and Japan(n=5754).In terms of national cooperation,China presented the highest centrality,its strongest bonds being to the United States and Japan.Among the 20 academic institutions with the most publications,ten came from China and the rest of Asia,though the University of Paris Cité,Public Assistance-Hospitals of Paris,and the National Institute of Health and Medical Research(INSERM)were the most prolific.As for individual contributions,Hisatomi H,Kaneko S,and Ide T were the three most prolific authors.Kaneko S ranked first by H-index,G-index,and overall publication count,while Zucman-Rossi J ranked first in citation count.The five most popular journals were the World Journal of Gastroenterology,Hepatology,Journal of Hepatology,Oncotarget,and Oncogene,while Nature Genetics,Hepatology,and Nature Reviews Disease Primers had the most citations.We extracted 2293 keywords from the publications,120 of which appeared more than ten times.The most frequent were HCC,telomerase and human telomerase reverse transcriptase(hTERT).Keywords such as mutational landscape,TERT promoter mutations,landscape,risk,and prognosis were among the most common issues in this field in the last three years and may be topics for research in the coming years.CONCLUSION Our bibliometric analysis provides a comprehensive overview of HCC/telomerase research and insights into promising upcoming research.

Anti-aging based on stem cell therapy:A scoping review

Stem cells are present in the tissues and organs and remain in a quiescent and undifferentiated state until it is physiologically necessary to produce new descendant cells.Due to their multipotency property,mesenchymal stem cells have attracted considerable attention worldwide due to their immunomodulation and therapeutic function in tissue regeneration.Stem cells secrete components such as paracrine factors,extracellular vesicles,and exosomes which have been shown to have anti-inflammatory,anti-aging,reconstruction and wound healing potentials in many in vitro and in vivo models.The pluripotency and immunomodulatory features of stem cells could potentially be an effective tool in cell therapy and tissue repair.Aging affects the capacity for self-renewal and differentiation of stem cells,decreasing the potential for regeneration and the loss of optimal functions in organisms over time.Current progress in the field of cellular therapy and regenerative medicine has facilitated the evolution of particular guidelines and quality control approaches,which eventually lead to clinical trials.Cell therapy could potentially be one of the most promising therapies to control aging due to the fact that single stem cell transplantation can regenerate or substitute the injured tissue.To understand the involvement of stem cells not only in tissue maintenance and disease but also in the control of aging it is important to know and identify their properties,functions,and regulation in vivo,which are addressed in this review.

Direct Correlation among Telomere Length, Cellular Aging, and Rejuvenation Effects of Honey Child Powder

Purpose: Telomere length (TL) is an indicator of age;however, hormonal influences complicate individual aging. It remains unclear whether TL shortening is a direct factor in both individual and cellular aging. Therefore, we examined the direct relationship between TL and cellular senescence at the cellular level. Methods: Telomerase activity, TL, and gene expression were measured in cultured human lung-, fetal-, and skin-derived fibroblasts, human skin keratinocytes, and telomerase reverse transcriptase (TERT) gene-immortalized cells using detection kits, Cawthon’s method, and reverse transcription-quantitative polymerase chain reaction, respectively. Novel substances that elongate telomeres were screened to confirm cell rejuvenation effects. Results: Long-term cell culture of TIG-1-20 normal human fibroblasts resulted in TL shortening, decreased division rate, and senescence progression, whereas in OUMS-36T-2 cells, TL elongation via TERT gene transfer increased the division rate, reduced endoplasmic reticulum stress, and upregulated genes associated with young individuals, indicating that cellular rejuvenation occurs via TL elongation. In addition, a honey child powder (HCP) extract was found through screening, and the HCP extract strongly suppressed the menin gene, resulting in increased telomerase activity and extended cell lifespan. Upon addition of the HCP extract to skin fibroblasts, gene expression of moisturizing components, including collagen, hyaluronic acid, and elastin, increased, and exhibited a rejuvenating effect with an increase in elastin amount. Conclusions: TL elongation or shortening is involved in cell proliferation rate and cellular aging, and TL elongation rejuvenates cells. In addition, HCP extract has a rejuvenating effect on cells and is expected to be a rejuvenating compound.

A Novel Replication-competent Adenovirus CNHK500 in the Treatment of Heptocellular Carcinoma In Vitro

Objective: To evaluate the therapeutic efficacy of replicative adenovirus CNHK500 in the treatment of hepatocellular carcinoma. Methods: Virus proliferation assay, cell viability assay and Western blot were performed to assess the selective replication and cytolysis of CNHK500 in telomerase positive liver cancer cells Hep3B, HepGII, SMMC7721 and in normal cells. Results: The replicative multiples of CNHK500 in HepGII, Hep3B and SMMC7221 after 96 h of virus proliferation were 52 000, 396 984.9 and 632 911.3 fold respectively, similar to those of wtAd5. However, CNHK500 demonstrated more significant attenuated replicative ability in normal cell lines than wtAd5. CNHK500 replicated only 3.1-100 fold at 96 h, while the wtAd5 still reached 3160-17 357 fold. CNHK500 could cause half of HepGII cells death within 7 days at MOI 2, in Hep3B cell lines the IC50 was as low as MOI 0.01, whereas the IC50 in BJ cell was as high as MOI 1000. CNHK500 E1A protein could only be detected in hepatocellular cancer cells but not in normal cells under normoxia. E1B protein could only be detected under hypoxia condition at a MOI of 1. Conclusion: CNHK500 can efficiently replicate in and kill liver cancer cells as well as wtAd5 do while it is severely attenuated in proliferation and cytolysis among normal cells. It would be a prominsing strategy for liver cancer tratment.

Inhibition of Telomerase with hTERT Antisense Increases Susceptibility of Leukemic Cells to CDDP-induced Apoptosis

Objective: To investigated the e?ect of inhibition of telomerase with hTERT antisense on leukemic cells (HL-60 and K562) to CDDP-induced apoptosis. Methods: Antisense phosphorothioate oligodeoxynucleotide (AS PS-ODN) was synthesized and puri?ed. Telomerase activity was detected by Telomerase PCR ELASA kit and cell apoptosis was observed by morphological method and determined by ?owcytometry. Results: AS PS-ODN could signi?cantly inhibit telomerase activity by down regulat- ing the hTERT expression, and increase the susceptibility of leukemic cells to CDDP-induced apoptosis. Conclusion: Inhibition of telomerase with hTERT antisense can increases the susceptibility of leukemic cells to CDDP-induced apoptosis.

Trichostatin A Induces Apoptosis by Inhibiting Telomerase Activity and Expression of Telomerase Reverse Transcriptase in HL-60 Cells

Aim To investigate the effects of trichostatin A （TSA） on telomerase activity and the expression of human telomerase reverse transcriptase （hTERT） during apoptosis in vitro and the mechanisms in HL-60 cells. Methods The proliferative activity of HL-60 cells was assessed by MTT assay. Cell apoptosis was analyzed by flow cytometry. Telomerase activity was examined by TRAP-ELISA. The expression of telomerase subunits was analyzed by RT-PCR. Results A time- and dose-dependent inhibition was detected in HL-60 cells treated with TSA. After treatment with 600 nmol· L^-1 TSA for 48 h, the apoptosis rate in HL-60 cells was 42. 6% and telomerase activity decreased 1.95 ± 0.25, 1.73 ± 0. 12, and 1.52 ± 0. 09 for 12, 24, and 48 h, respectively. The expression of hTERTmRNA decreased. No significant changes were observed in the expression of hTRmRNA and hTPI mRNA. Condusion TSA inhibits telomerase activity and induces apoptosis in HL-60 cells. The underlying mechanism may be related to the down-regulation of hTERT transcription.

Inhibitory Effect of Oridonin on the Proliferation of NB4 Cells and Its Mechanism

Objective: To investigate the anti-proliferation e?ect of oridonin on leukemic NB4 cells and its mechanism. Methods: NB4 cells in culture medium in vitro were given di?erent concentrations of oridonin. The inhibitory rate of the cells were measured by MTT assay, cell apoptotic rate was detected by ?ow cytometry(FCM), morphology of cell apoptosis was observed by hoechst 33258 ?uorescence staining , and the activity of telomerase was detected using TRAP-PCR-ELISA before and after apoptosis occurred. Results: Oridonin (over 8 μmol/L) could decrease the telomerase activity, inhibit the growth of NB4 cells and induce apoptosis signi?cantly in a time- and dose-dependent manner. Marked morphological changes of cell apoptosis were observed by hoechst 33258 ?uorescence staining especially after the cells treated by oridonin for 48–60 h. Conclusion: Oridonin could inhibit the proliferation and induce the apoptosis of NB4 cells in vitro. One of the mechanisms may be the decrease of the telomerase activity of NB4 cells.

Telomerase Activity in Human Renal Cell Carcinoma with Reference to Clinicopathologic Features

Objective: To study the telomerase activity in human renal cell carcinoma and to evaluate the correlation with the clinicobiologic features of the neogrowth.Methods: The telomerase activity was studied by means of a modified telomeric repeat amplification protocol (TRAP) in 32 renal cell carcinoma tissues, 32 normal renal tissues and 32 paracancer tissues and its correlation with the clinicopathologic features of the tumor was evaluated.Results: Telomerase activity was strongly positive in 17, positive in 12 and negative in 3 cases of renal cell carcinoma tissues, the total positive rate being 91%. Telomerase activity was weakly positive (6%) in only 2 out of 32 samples of normal renal cortex tissues and positive in 6 paracancer tissues (19%), the difference was conspicuous (P<0.01).Conclusion: The positive rate of telomerase activity was significantly higher in renal cell carcinoma tissues and might serve as a prognostic marker for estimating the biologic characteristics of renal cell carcinoma.

Expression of Telomerase Activity in Gastric Cancer Cells

Objective To study the relationship between telomerase activity and biological behavior in human gastric cancer cells and appraise the clinical significance of detecting telomerase activity.Methods The telomerase activity in 47 gastric cancer tissue samples , their matched normal tissues, 7 gastric ulcer and 2 gastric cancer cell lines was detected using a PCR-based non-radioisotopic telomeric repeat amplification protocol (TRAP) assay. Results None of the 47 samples from normal gastric tissues expressed telomerase activity. The 41 of 47 cases of gastric cancer presented telomerase activity with an 87.2 % positive rate (P < 0.001). 2/2 gastric cancer cell lines and 0/7 gastric ulcer line were also positive for telomerase activity. The activity of telomerase was associated with the pathological differentiation of gastric cancer. Conclusion Telomerase activity may be related to the biological behavior of gastric cancer and can help in assessing the malignant potential of gastric cancer. Telomerase activity will be a good diagnostic marker for the detection of gastric cancer.

Effects of Artificially Designed shRNA of chTR on Telomerase Activity

[Objective] The study was to investigate the inhibitory effects of shRNA of chTR on telomerase activity in MDCC-MSB1 cells. [Method] The artificially designed and synthesized short hairpin RNA(shRNA) of chicken telomerase RNA(chTR) was used to construct the expression vector that was subsequently transfected to MDCC-MSB1 cells, then the telomerase activity of these cells measured by improved TRAP method. [Result] Compared to the CK group, the telomerase activities in the treatment cells showed no obvious decrease in 24 h after transfection, while decreased remarkably 48 h later, among which in the cells transfected by interference vector pSi-chTR-sh1 presented the most obvious inhibitory effects. [Conclusion] This vector could effectively inhibit the telomerase activities in MDCC-MSB1 cells.

端粒酶和血管内皮生长因子C在鼻腔鼻窦鳞状细胞癌组织中的表达及意义

近年来鼻腔鼻窦原发肿瘤有增长趋势，为进一步了解鼻腔鼻窦肿瘤临『末及病理特点，我们初步应用免疫组织化学技术检测了鼻腔鼻窦鳞状细胞癌中端粒酶和血管内皮生长因子C（vascular endothelial growth factorC,VEGF-c）的表达情况。

Telomerase activity in gastric cancer and its clinical implications

AIM To study the telomerase expression in gastric carcinoma and its clinical implications. METHODS Telomerase activity was examined in gastric cancer and corresponding normal tissues using a modified TRAP (telomeric repeat amplification protocol) assay (TRAP eze) in tissue samples from 94 gastric carcinomas and 58 normal tissues, 12 gastric adenomas and 9 gastric ulcer lesions. RESULTS Telomerase activity was present in 81 of the 94 (86 2%) gastric cancer tissues, whereas no telomerase activity was detected in any normal tissues. The incidence of telomerase activity in gastric cancer tissues was unrelated to the tumor diameter, histological grade, tumor invasion in depth, lymph node metastasis and TNM stage. CONCLUSION Telomerase plays an important role in carcinogenesis and progression of gastric cancer, and it is suggested to be a useful tumor marker.

The prognostic molecular markers in hepatocellular carcinoma

The prognosis of hepatocellular carcinoma (HCC) still remains dismal, although many advances in its clinical study have been made. It is important for tumor control to identify the factors that predispose patients to death. With new discoveries in cancer biology, the pathological and biological prognostic factors of HCC have been studied quite extensively. Analyzing molecular markers (biomarkers) with prognostic significance is a complementary method. A large number of molecular factors have been shown to associate with the invasiveness of HCC, and have potential prognostic significance. One important aspect is the analysis of molecular markers for the cellular malignancy phenotype. These include alterations in DNA ploidy, cellular proliferation markers (PCNA, Ki-67, Mcm2, MIB1, MIA, and CSE1L/CAS protein), nuclear morphology, the p53 gene and its related molecule MD M2, other cell cycle regulators (cyclin A, cyclin D, cyclin E, cdc2, p27, p73), oncogenes and their receptors (such as ras, c-myc, c-fms, HGF, c-met, and erb-B receptor family members), apoptosis related factors (Fas and FasL), as well as telomerase activity. Another important aspect is the analysis of molecular markers involved in the process of cancer invasion and metastasis. Adhesion molecules (E-cadherin, catenins, serum intercellular adhesion molecule-1, CD44 variants), proteinases involved in the degradation of extracellular matrix (MMP-2, MMP-9, uPA, uPAR, PAI), as well as other molecules have been regarded as biomarkers for the malignant phenotype of HCC, and are related to prognosis and therapeutic outcomes. Tumor angiogenesis is critical to both the growth and metastasis of cancers including HCC, and has drawn much attention in recent years. Many angiogenesis-related markers, such as vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), platelet-derived endothelial cell growth factor (PD-ECGF), thrombospondin (TSP), angiogenin, pleiotrophin, and endostatin (ES) levels, as well as intratumor microvessel density (MVD) have been evaluated and found to be of prognostic significance. Body fluid (particularly blood and urinary) testing for biomarkers is easily accessible and useful in clinical patients. The prognostic significance of circulating DNA in plasma or serum, and its genetic alterations in HCC are other important trends. More attention should be paid to these two areas in future. As the progress of the human genome project advances, so does a clearer understanding of tumor biology, and more and more new prognostic markers with high sensitivity and specificity will be found and used in clinical assays. However, the combination of some items, i.e., the pathological features and some biomarkers mentioned above, seems to be more practical for now.

Effect of VEGF,P53 and telomerase on angiogenesis of gastric carcinoma tissue

Objective:To investigate the effect of vascular endothelial growth factor(VEGF),P53 and telomerase on angiogenesis in gastric carcinoma tissue.Methods:A total of 95 surgical resection samples of gastric cancer tissue after pathological diagnosis are collected to observe the VEGF,P53 and telomerase expression using immunohistochemical methods.Relationship between their expression and its influence on angiogenesis in gastric carcinoma tissue were analyzed.Results:Microvascular density(MVD)and the expression of VEGF,P53 and telomerase were positively correlated.Expression of VEGF and P53 protein were related to tumor type and lymph metastasis,and also a correlation was observed between P53 and VEGF.The telomerase expression had no correlation with VEGF,and P53.Conclusions:VEGF angiogenesis has a angiogenesis promoting effect on gastric cancer tissue development and plays an important role in tumor generation and metastasis.Mutant P53 promotes the tumor angiogenesis generation by adjusting VEGF.Telomerase has a certain role in promoting activity of angiogenesis through different way rather than P53.

Study of the expressions of p53 and bcl-2 genes, the telomerase activity and apoptosis in GIST patients

AIM： To explore the relationship between clinicobiological behavior and the expression levels of telomerase activity, apoptosis, p53 gene and bcl-2 gene in gastrointestinal stromal tumors （GISTs）. METHODS： The intensity of telomerase activity, apoptosis, p53 and bcl-2 expression in GISTs were detected by telomeric repeat amplification protocol, in situ end-labeling technique, and immunohistochemistry, respectively. RESULTS： The positive rates of telomerase activity of malignant GIST, potential malignant GIST and benign GIST were 85% （17/20）, 22.8% （2/9） and 0 （0/9）, respectively. The apoptosis indices of malignant GIST, potential malignant GIST, and benign GIST were 11.7±5.4, 30.2±5.6 and 45.2 ±7.2, respectively. The intensity of telomerase activity and apoptosis were related to the biological characteristics of GISTs （85% vs 22.8%, 0, 0; P 〈 0.01 or 11.7±5.4 vs 30.2±5.6, 45.2±7.2, 72.1±9.3; P 〈 0.05）. The intensity of telomerase activity was negatively correlated with cellular apoptosis （22.9±8.4 vs 9.5±5.7, P 〈 0.01）. The intensity of telomerase activity was positively correlated with/753, bcl-2 expression （40.0% vs 78.9%, 40.0% vs 84.2%; P 〈 0.05）. CONCLUSION： The detection of telomerase activity, apoptosis and its control genes in GIST will be helpful for the discrimination of the malignant and benign GIST and evaluation of the prognosis.

Combination of telomerase antisense oligonucleotides simultaneously targeting hTR and hTERT produces synergism of inhibition of telomerase activity and growth in human colon cancer cell line

AIM: To investigate synergism of inhibition of telomerase activity and proliferation of human colon cancer cells by combination of telomerase antisense oligonucleotides (ASODNs) simultaneously targeting human telomerase RNA (hTR) and human telomerase reverse transcriptase (hTERT) in vitro. METHODS: ASODN of hTR and ASODN of hTERT were transfected into human colon cancer SW480 cells by liposomal transfection reagents. Telomerase activity of SW480 cells was examined using telomeric repeat amplification protocol (TRAP)-enzyme-linked immunosorbent assay (PCR-ELISA). Proliferation activity of SW480 cells was tested by methyl thiazolyl tetrazolium assay. Apoptosis and cell cycle were analyzed by flow cytometry. RESULTS: The telomerase activity and cell survival rate in SW480 cells transfected with 0.2 μmol/L of ASODN of hTR or ASODN of hTERT for 24-72 h were significantly decreased in a time-dependent manner compared with those after treatment with sense oligonucleotides and untreated (telomerase activity: 24 h, 73%, 74% vs99%, 98%; 48 h, 61%, 55% vs98%, 99%; 72 h, 41%, 37% vs 99%, 97%; P<0.01; cell survival rate: 24 h, 88%, 86% vs594%, 98%; 48 h, 49%, 47% vs94%, 97%; 72 h, 44%, 42% vs92%, 96%; P<0.01). Moreover, the telomerase activity and the cell survival rate in SW480 cells treated by the combination of telomerase anti-hTR and anti-hTERT were more significantly suppressed than single anti-hTR or anti-hTERT (telomerase activity: 24 h, 59% vs 73%, 74%; 48 h, 43% vs61%, 55%; 72 h, 18% vs41%, 37%; P<0.01; cell survival rate: 24 h, 64% vs88%, 86%; 48 h, 37% vs49%, 47%; 72 h, 25% vs44%, 42%; P<0.01). Meanwhile, the apoptosis rates in the combination group were markedly increased compared with those in the single group (24 h, 18.0% vs7.2%, 7.4%; 48 h, 23.0% vs13.0%, 14.0%; 72 h, 28.6% vs 13.2%, 13.75; P<0.01). Cells in combination group were arrested at G0/G1 phase. CONCLUSION: Telomerase anti-hRT and anti-hTERT suppress telomerase activity, and inhibit growth of human colon cancer cells probably via induction of apoptosis and retardation of cell cycle. Additionally, combined use of telomerase ASODNs targeting both hTR and hTERT yields synergistic action selective for human colon cancer.

Telomere and telomerase in chronic liver disease and hepatocarcinoma

The pathogenesis of liver cirrhosis is not completely elucidated.Although in the majority of patients,the risk factors may be identified in B and C viral hepatitis,alcohol intake,drugs or fatty liver disease,there is a small percentage of patients with no apparent risk factors.In addition,the evolution of chronic liver disease is highly heterogeneous from one patient to another.Among patient with identical risk factors,some rapidly progress to cirrhosis and hepatocellular carcinoma(HCC)whereas others have a benign course.Therefore,a genetic predisposition may contribute to the development of cirrhosis and HCC.Evidence supporting the role of genetic factors as a risk for cirrhosis has been accumulating during the past years.In addition to the results from epidemiological studies,polymorphisms studies and data on twins,the concept of telomere shortening as a genetic risk factor for chronic liver disease and HCC has been proposed.Here we review the literature on telomerase mutations,telomere shortening and liver disease including hepatocellular carcinoma.

Using a non-radioisotopic, quantitative TRAP-based method detecting telomerase activities in human hepatoma cells

A non-radioisotopic, quantitative TRAP-based telomerase activity assay was established mainly by using SYBR Green-I staining instead of radioisotope. Comparing with conventional radioisotope based method, it was better in reproducibility and accuracy. Using this method, we found telomerase activities were absent in normal human liver cells, while detected in ail of four human hepatoma cell lines (BEL-7404, SMMC-7721, QGY-7903 and HCCM) without significant differences.

Inhibitory Effects of Selenium on Telomerase Activity and hTERT Expression in Cadmium-transformed 16HBE Cells

To investigate the effects of sodium selenite on telomerase activity and expression of hTERT mRNA in cadmium-transformed 16HBE cells. Methods Telomerase activity and expression of genes were measured after cultured cadmium-transformed 16HBE cells were exposed to sodium selenite at different doses （0.625, 1.25, 2.50, 5.00 pmol/L） for 24 hours. Results Selenium decreased telomerase activity in cadmium-transformed 16HBE cells. There existed an obvious dose-effect relationship between the selenium concentration and these changes. The expression of hTERT and c-myc mRNA also decreased but the expression of madl mRNA increased after exposure to selenium for 24 hours. No difference was found in expression of hTRF1 and hTRF2 mRNA after incubated with sodium selenite for 24 hours, compared with control group. Conclusion Selenium inhibits telomerase activity by decreasing hTERT and c-myc mRNA expression and increasing madl mRNA expression in cadmium-transformed 16HBE cells and selenium concentration is significantly correlated with these changes.