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We attempted to clarify myocardial telomere dynamics using samples from 530 autopsied patients using Southern blot analysis. Overall regression analysis demonstrated yearly telomere reduction rate of 20 base pairs in the myocardium. There was a significant correlation between myocardial telomere and aging. Moreover, regression analyses of telomere and heart weight yielded a telomere reduction rate of 3 base pairs per gram, and a small but significant correlation between telomere reduction and heart weight was demonstrated. Hearts of autopsied patients who had died of heart disease were significantly heavier than those of patients who had died of cancer or other diseases, and heart disease was significantly more correlated with myocardial telomere shortening than cancer or other diseases. Here we show that telomeres in myocardial tissue become shortened with aging and heart disease, and that heart disease was associated with a gain of heart weight and telomere shortening in the myocardium. [ABSTRACT FROM AUTHOR]

Abstract Despite a number of in vitro studies of transthyretin (TTR) amyloidogenesis the early stage of in vivo amyloidogenesis in the human heart is largely unknown. A heart with a mild degree of cardiac amyloidosis removed from a 90-year old woman at autopsy was selected for analysis. The genotype of the TTR was the wild type. An immunohistochemical study with anti-TTR antibody was performed on serial paraffin sections, and 17 TTR-positive lesions less than 50 µm in diameter consisting of 13 interstitial and 4 vascular lesions were identified. The early interstitial lesions start as thick membranous deposits between interfacing myocardial cells. They are Congophilic with green birefringence and positive for apolipoprotein E but negative for amyloid P component. The TTR-positive amyloid extends along intercellular spaces and becomes larger, involving several myocardial fibers. The media is the initial site of arteriolar involvement. According to the in vitro studies of amyloid fibrillogenesis, the most critical step is formation of the nucleus under supersaturated conditions. The supersaturated conditions are speculated to be achieved by binding to proteoglycans or lipid membranes. Our results indicate that the basement membrane of myocardial cells is the initial site of amyloid deposition, providing a suitable place for concentration of TTR. [ABSTRACT FROM AUTHOR]

Haem oxygenase (HO)-1, a rate-limiting enzyme in the degradation of haem, is increased in Alzheimer's disease and in inflammations such as AA amyloidosis. However, the specific association of HO-1 is poorly understood in AA amyloidosis. In this study, we designed the experiment to reveal the contribution and association of HO-1 in the spleen during experimental murine AA amyloidosis. Experimental murine AA amyloidosis was induced with injection of an emulsion consisting of Freund's complete adjuvant and Mycobacterium butyricum. The serum amyloid A level was highest on day 3. The distribution of cells containing iron, indicating an increase of HO-1 in the red pulp, was detected with Berlin blue staining. AA amyloid formation was immunohistochemically detected as a marker by chondroitin sulphate proteoglycan (CSPG), one of the components of AA amyloid fibrils. Immunolocalizations of HO-1 and CSPG indicated a conspicuous increase and scattering of positive cells in the red pulp of the spleen. Double positive cells were not detected. On day 7, amyloid deposition was detected with Congo red staining in the extracellular spaces in the marginal zone of the white pulp in the spleen and HO-1-positive cells accumulated near the amyloid deposition area. CSPG was detected within the cells and also localized in the amyloid deposition area. CSPG was still not localized in the HO-1-positive cells. Double positive cells of HO-1 and CSPG were localized in the red pulp and in the amyloid deposition area on day 14. X-ray microanalysis indicated the existence of iron in the electron-dense bodies of fibroblasts in the amyloid deposition areas. The fibroblasts extended amyloid fibrils into the extracellular spaces of the marginal zone. These results suggest that HO-1-positive fibroblasts, but not HO-1-positive macrophages, are associated with the late stage of amyloid fibril formation. [ABSTRACT FROM AUTHOR]

A large body of evidence supports a key role for telomere dysfunction in carcinogenesis due to the induction of chromosomal instability. To study telomere shortening in precancerous pancreatic lesions, we measured telomere lengths using quantitative fluorescence in situ hybridization in the normal pancreatic duct epithelium, pancreatic intraepithelial neoplasias (PanINs), and cancers. The materials employed included surgically resected pancreatic specimens without cancer (n = 33) and with invasive ductal carcinoma (n = 36), as well as control autopsy cases (n = 150). In comparison with normal ducts, telomere length was decreased in PanIN-1, −2 and −3 and cancer. Furthermore, telomeres were shorter in cancer than in PanIN-1 and −2. Telomere length in cancer was not associated with histological type, lesion location, or cancer stage. PanINs with or without cancer showed similar telomere lengths. The incidences of atypical mitosis and anaphase bridges, which are morphological characteristics of chromosomal instability, were negatively correlated with telomere length. The telomeres in normal duct epithelium became shorter with aging, and those in PanINs or cancers were shorter than in age-matched controls, suggesting that telomere shortening occurs even when histological changes are absent. Our data strongly suggest that telomere shortening occurs in the early stages of pancreatic carcinogenesis and progresses with precancerous development. Telomere shortening and chromosomal instability in the duct epithelium might be associated with carcinogenesis of the pancreas. Determination of telomere length in pancreatic ductal lesions may be valuable for accurate detection and risk assessment of pancreatic cancer. [ABSTRACT FROM AUTHOR]

In the original publication of this article, Fig. 5 was published with incorrect color of the approximate lines of CBD and Control. [ABSTRACT FROM AUTHOR]

Aim: The telomere is a structure present at the ends of chromosomes, and is known to shorten with aging and successive rounds of cell division. However, very little is known about telomere attrition in post‐mitotic cells, such as neurons. Methods: Using our originally developed quantitative fluorescence in situ hybridization method, we analyzed age‐dependent alterations of telomere length in three types of cells in the human cerebrum: neurons and glial cells in both the gray and white matter. Results: In adults, telomeres were significantly longer in neurons than in glial cells, whereas in infants, telomere lengths did not differ among the three cell types. No aging‐related telomere attrition was evident in neurons. However, the telomeres of glial cells were shorter in older individuals than in younger individuals, and attrition was more rapid in the white matter than in the gray matter. Conclusions: The present results suggest that the telomeres of neurons remain stable throughout life, whereas telomeres in white matter glial cells become significantly shorter with age. Examination of adults showed no significant correlation between telomere length and age in the three cell types. Although the present study was cross‐sectional, the results suggest that telomere shortening before adolescence contributes to the significant decrease of telomere length in white matter glial cells. The present findings in normal cerebral tissues will be informative for future studies of telomere stability in the diseased brain. Geriatr Gerontol Int 2018; 18: 1507–1512. [ABSTRACT FROM AUTHOR]

Background: Congenital biliary dilatation (CBD) is a congenital malformation involving both dilatation of the extrahepatic bile duct and pancreaticobiliary maljunction. Persistent reflux of pancreatic juice injures the biliary tract mucosa, resulting in chronic inflammation and higher rates of carcinogenesis in the biliary tract, including the gallbladder. Telomeres are repetitive DNA sequences located at the ends of chromosomes. Chromosomal instability due to telomere dysfunction plays an important role in the carcinogenesis of many organs. This study was performed to determine whether excessive shortening of telomeres occurs in the gallbladder mucosa of patients with CBD.Methods: Resected gallbladders were obtained from 17 patients with CBD, ten patients with cholecystolithiasis without pancreatic juice reflux, and 17 patients with normal gallbladders (controls) (median age of each group of patients: 37, 50, and 53 years, respectively). The telomere lengths of the gallbladder epithelium were measured by quantitative fluorescence in situ hybridization using tissue sections, and the normalized telomere-to-centromere ratio (NTCR) was calculated.Results: The NTCRs in the CBD, cholecystolithiasis, and control groups were 1.24 [interquartile range (IQR) 1.125-1.52], 1.96 (IQR 1.56-2.295), and 1.77 (IQR 1.48-2.53), respectively. The NTCR in the CBD group was significantly smaller than that in the cholecystolithiasis and control groups (p = 0.003 and 0.004, respectively), even in young patients.Conclusions: Our findings indicate that telomere shortening in the gallbladder mucosa plays an important role in the process of carcinogenesis in patients with CBD. These results support the recommendation of established guidelines for prophylactic surgery in patients with CBD because CBD is a premalignant condition with excessive telomere shortening. [ABSTRACT FROM AUTHOR]

Cancer stem cells (CSCs), which are pluripotent and self-renewable, contribute to the initiation and metastasis of cancer, and are responsible for resistance to chemotherapy and radiation. Pancreatic ductal adenocarcinoma (PDAC) is an aggressive type of cancer that is associated with a high incidence of distant metastasis and recurrence. Sphere formation reveals cell proliferation under nonadherent conditions and is commonly used to identify CSCs; measurements of the number, area and volume of the spheres are used to estimate stemness of PDAC cells. However, detailed morphological analysis of such spheres has not been performed. The aim of the present study was to examine the morphology of spheres isolated from PANC-1 human pancreatic cancer cells via scanning electron microscopy (SEM) and transmission electron microscopy (TEM). PANC-1 cells formed round to irregular oblong spheres within 1 week following seeding in ultra-low-attachment plates. These spheres exhibited higher levels of expression of CSC markers, including nestin, sex determining region Y-box 2, and CD44 containing variant exon 9, compared with adherent cells. SEM analysis revealed that the spheres exhibited a grape-like appearance, harboring cancer cells with smooth or rough surfaces. Similarly, TEM analysis detected cancer cells with varying surface types within the spheres: Those with smooth surfaces, irregular large protrusions, protrusions and a small number of microvilli, and those with many microvilli throughout the entire cell surface. These morphological differences among cancer cells may be indicative of different stages in the differentiation process, from CSCs to non-CSCs, within the spheres. [ABSTRACT FROM AUTHOR]

The article offers the authors' views on operational tolerance (OT) and its association with recipient age in living donor liver transplantation (LDLT). They mention the essential factor of donor age on the telomere length sustainability in hepatocytes. They also note the significance of protocol liver biopsy (PLB) in the evaluation of histopathology.

We have reported telomere attrition in β and α cells of the pancreas in elderly patients with type 2 diabetes, but it has not been explored how the telomere lengths of these islet cells change according to age in normal subjects. To examine the telomere lengths of β and α cells in individuals without diabetes across a wide range of ages, we conducted measurement of the telomere lengths of human pancreatic β and α cells obtained from 104 autopsied subjects without diabetes ranging in age from 0 to 100 years. As an index of telomere lengths, the normalized telomere-centromere ratio (NTCR) was determined for β (NTCRβ) and α (NTCRα) cells by quantitative fluorescence in situ hybridization (Q-FISH). We found NTCRβ and NTCRα showed almost the same levels and both decreased according to age (p < 0.001 for both). NTCRs decreased more rapidly with age and were more widely distributed (p = 0.036 for NTCRβ, p < 0.001 for NTCRα) in subjects under 18 years of age than in subjects over 18 years. There was a positive correlation between NTCRβ and NTCRα only among adult subjects (p < 0.001). In conclusion, the telomeres of β and α cells become shortened with normal aging process. [ABSTRACT FROM AUTHOR]

Accumulated data have shown that most human somatic cells or tissues show irreversible telomere shortening with age, and that there are strong associations between telomere attrition and aging-related diseases, including cancers, diabetes and cognitive disorders. Although it has been largely accepted that telomere attrition is one of the major causes of aging-related disorders, critical aspects of telomere biology remain unresolved, especially the lack of standardized methodology for quantification of telomere length. Another frustrating issue is that no potentially promising methods for safe prevention of telomere erosion, or for telomere elongation, have been devised. Here, we review several methods for quantification of telomere length currently utilized worldwide, considering their advantages and drawbacks. We also summarize the results of our recent studies of human cells and tissues, mainly using quantitative fluorescence in situ hybridization and Southern blotting, including those derived from patients with progeria-prone Werner syndrome and trisomy 21, and several strains of induced pluripotent stem cells. We discuss the possible merits of using telomere shortness as an indicator, or a new marker, for diagnosis of precancerous states and aging-related disorders. In addition, we describe newly found factors that are thought to impact telomere dynamics, providing a new avenue for examining the unsolved issues related to telomere restoration and maintenance. Geriatr Gerontol Int 2016; 16 (Suppl. 1): 30-42. [ABSTRACT FROM AUTHOR]

Along with the increasing need for living-donor liver transplantation (LDLT), the issue of organ shortage has become a serious problem. Therefore, the use of organs from elderly donors has been increasing. While the short-term results of LDLT have greatly improved, problems affecting the long-term outcome of transplant patients remain unsolved. Furthermore, since contradictory data have been reported with regard to the relationship between donor age and LT/LDLT outcome, the question of whether the use of elderly donors influences the long-term outcome of a graft after LT/LDLT remains unsettled. To address whether hepatocyte telomere length reflects the outcome of LDLT, we analyzed the telomere lengths of hepatocytes in informative biopsy samples from 12 paired donors and recipients (grafts) of pediatric LDLT more than 5 years after adult-to-child LDLT because of primary biliary atresia, using quantitative fluorescence in situ hybridization (Q-FISH). The telomere lengths in the paired samples showed a robust relationship between the donor and grafted hepatocytes (r = 0.765, p = 0.0038), demonstrating the feasibility of our Q-FISH method for cell-specific evaluation. While 8 pairs showed no significant difference between the telomere lengths for the donor and the recipient, the other 4 pairs showed significantly shorter telomeres in the recipient than in the donor. Multiple regression analysis revealed that the donors in the latter group were older than those in the former (p = 0.001). Despite the small number of subjects, this pilot study indicates that donor age is a crucial factor affecting telomere length sustainability in hepatocytes after pediatric LDLT, and that the telomeres in grafted livers may be elongated somewhat longer when the grafts are immunologically well controlled. [ABSTRACT FROM AUTHOR]

Many studies have demonstrated the association between telomere length in mitotic cells and aging, but the relationship between telomere length in post-mitotic cells and aging in human populations has remained largely unclear. In this study, we assessed the dynamics of telomere length (terminal restriction fragment length-TRF) in normal pituitary glands obtained at autopsy from 65 patients aged between 0 and 100 years. The initial length (the value for neonatal pituitary) was 14.2 ± 1.1 kbp (mean of the median TRF value ± SD), being the longest among those for other systemic organs. The annual telomere reduction rates for pituitary (21 bp) were equivalent to or below the lowest values for various cells and tissues. Hence, comparison of TRF lengths among organs revealed that the pituitary gland retained longer telomeres than most other organs throughout life. The median TRF value showed a significant decrease between newborns and the 61-75 age group, but then leveled off or increased slightly in advanced age groups. These data indicate that innate telomere length is highly conserved in the pituitary and suggest that pituitary telomere length provides a good basic indicator for studies of telomere biology. [ABSTRACT FROM AUTHOR]

Chromosomal and genomic instability due to telomere dysfunction is known to play an important role in carcinogenesis. To study telomere dysfunction in the surrounding background epithelium of squamous cell carcinoma in situ (CIS) of the oesophagus, we measured telomere lengths of basal and parabasal cells of epithelia with and without CIS using quantitative fluorescence in situ hybridization (Q-FISH) and our original software, Tissue Telo. Additionally, we assessed histological inflammation and the anaphase bridge index. In non-cancerous epithelium, telomeres in basal cells were significantly longer than those in parabasal cells, whereas CIS showed a homogeneous telomere pattern in the basal and parabasal cells. Telomeres in basal and parabasal cells were significantly shorter in the background with CIS than in epithelium from age-matched normal controls. Significant negative correlation was observed between the normalized telomere : centromere ratio (reflected telomere length) and the anaphase bridge index in non-cancerous epithelia from both normal controls and the CIS background with no histological inflammation. These findings indicate that tissue stem cells may be located among basal cells, and that telomere length distribution in component cell types differs between CIS and non-cancerous epithelium. We have demonstrated conclusively that oesophageal CIS arises from epithelium with short telomeres and chromosomal instability in the absence of histological inflammation. Copyright © 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Objective. Telomere shortening is thought to be associated with genetic instability. The purpose of this study was to measure telomere length in a series of Barrett's adenocarcinomas (BAs), focusing on the telomere/centromere fluorescent intensity ratio (TCR) with tissue quantitative fluorescent in situ hybridization (Q-FISH). Material and methods. A total of 11 cases of BA were evaluated for upper esophagus (UE), lower esophagus (LE), Barrett's mucosa (BM), BA, and gastric cardiac mucosa (GC). Q-FISH was performed using two kinds of peptide nucleic acid probe, specific for telomeres and centromeres. The sections were analyzed with a CCD camera and original software (Tissue Telo) for measuring TCR. In addition, Laser Capture Microdissection and GeneScan were implemented for evaluation of genetic instability. Results. The TCR values in BM and, to a lesser extent, BA were significantly lower than those in the other tissues, particularly in heterozygosity (LOH)-positive cases. However, no significant difference was evident between microsatellite instability (MSI)-positive and -negative groups. Conclusions. In our study of BA series, telomere length appeared to change with the degree of histological atypia, with decreases linked to LOH. [ABSTRACT FROM AUTHOR]

Objective: We have developed a novel method for evaluating telomere length in four different cell types in non-cancerous and cancerous mucosal tissue from 15 cases of squamous cell carcinoma of the esophagus using tissue quantitative fluorescence in situ hybridization (Q-FISH). We hypothesized that the very rapid cell proliferation observed in esophageal squamous cell carcinomas might accelerate the telomere shortening and chromosomal instability associated with carcinogenesis. Methods: Tissue Q-FISH and the telomere to centromere intensity ratio (TCR) were used to compare telomere shortening in tissue sections taken from esophageal squamous cell carcinomas and adjacent non-cancerous esophageal tissues. Results: The peak percentage of TCR was <1 for esophageal squamous carcinoma cells and >1 for the non-cancerous esophageal cell types. Basal layer cells had the longest telomeres in comparison with prickle, cancer, and stromal cells, and strongly expressed hTERT, cytokeratin 14 and CD49f, but not MIB-1. Conclusion: These results suggest the presence of stem cells in the basal layer of the esophagus. Esophageal squamous cell carcinomas also display anaphase bridges, evidencing chromosomal instability.In conclusion, our TCR method can be used to distinguish between benign and malignant tissue in esophageal lesions. In order to apply this approach clinically to individual cases, further studies are in progress. Copyright © 2006 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Summary background and objective Telomerase activity is present in most malignant tumours and might provide a mechanism for unlimited replication of neoplastic cells. Expression of the gene encoding human telomerase reverse transcriptase (hTERT), the telomerase catalytic subunit gene, is associated with telomerase activity, and it is overexpressed in most parathyroid carcinomas. We have therefore studied hTERT expression in parathyroid tumours. methods In the present study, we assayed telomerase activity by the telomeric repeat amplification protocol (TRAP) and used in situ hybridization (ISH) to study hTERT mRNA expression in four parathyroid metastatic cancers, six adenomas and two parathyroid hyperplasia tissue specimens. results Telomerase activity was detected by the TRAP assay in all of the parathyroid cancers (100%) but in none of the eight parathyroid benign lesions. hTERT gene expression was detected in the nuclei of the parathyroid cancer cells in all of the lesions (100%) but in none of the eight parathyroid benign lesions. conclusions Our results demonstrate a correlation between telomerase activity and hTERT gene expression measured by ISH (P < 0·001) in parathyroid tumours. We succeeded in very clearly and sensitively demonstrating hTERT mRNA in parathyroid tissues by using a new probe. [ABSTRACT FROM AUTHOR]

Progressive telomere shortening with aging was studied using normal esophageal mucosal specimens from 177 human subjects aged between 0 and 102 years (yrs). We observed age-related shortening of the telomere, at a rate of 60 base pairs (bp) per year (yr). The mean telomere length of 12 neonates was 15.2 kilobase pairs (kbp) and that of 2 centenarians was 9.3 kbp. Mean (±SD) telomere lengths were 14.9±1.3, 14.0±1.8, 10.1±3.7, 10.4±3.3 and 9.5±3.1 kbp for the age groups less than 2 yrs, 2–20 yrs, 21–60 yrs, 61–80 yrs and 81–102 yrs, respectively. The variation in telomere length among individuals in the same age group was greater for the 3 older groups than for the 2 younger groups, as shown by the SDs. Furthermore, older individuals had greater telomere length variation than younger individuals, based on the lengths of DNA digested smears. Although the telomere length decreased significantly with aging at the rate of 60 bp per yr, differences in the mean telomere lengths between the 3 older age groups were not significant. Rapid shortening occurred in the young generations and there was no further substantial decrease in the esophageal mucosa after 60 yrs of age. Compared to the very rapid renewal rate of the esophageal epithelial cells, the annual reduction rate in telomere length was very low. These findings support the hypothesis that germ cells in the esophageal epithelium have a mechanism to lengthen telomeres. [ABSTRACT FROM AUTHOR]