scholarly journals Lung alveolar integrity is compromised by telomere shortening in telomerase-null mice

2009 ◽  
Vol 296 (1) ◽  
pp. L57-L70 ◽  
Author(s):  
Jooeun Lee ◽  
Raghava Reddy ◽  
Lora Barsky ◽  
Jessica Scholes ◽  
Hui Chen ◽  
...  
Keyword(s):  
Absolute Number ◽  
Cell Number ◽  
Telomere Maintenance ◽  
Alveolar Wall ◽  
Wild Type ◽  
Telomere Shortening ◽  
Alveolar Epithelial ◽  
Epithelial Compartment ◽  
Distal Lung ◽  
Dna Strand

Shortened telomeres are a normal consequence of cell division. However, telomere shortening past a critical point results in cellular senescence and death. To determine the effect of telomere shortening on lung, four generations of B6.Cg- Terc tm1Rdp mice, null for the terc component of telomerase, the holoenzyme that maintains telomeres, were bred and analyzed. Generational inbreeding of terc−/− mice caused sequential shortening of telomeres. Lung histology from the generation with the shortest telomeres ( terc−/− F4) showed alveolar wall thinning and increased alveolar size. Morphometric analysis confirmed a significant increase in mean linear intercept (MLI). terc−/− F4 lung showed normal elastin deposition but had significantly decreased collagen content. Both airway and alveolar epithelial type 1 cells (AEC1) appeared normal by immunohistochemistry, and the percentage of alveolar epithelial type 2 cells (AEC2) per total cell number was similar to wild type. However, because of a decrease in distal lung cellularity, the absolute number of AEC2 in terc−/− F4 lung was significantly reduced. In contrast to wild type, terc−/− F4 distal lung epithelium from normoxia-maintained mice exhibited DNA damage by terminal deoxynucleotidyltransferase (TdT)-mediated dUTP nick end labeling (TUNEL) and 8-oxoguanine immunohistochemistry. Western blotting of freshly isolated AEC2 lysates for stress signaling kinases confirmed that the stress-activated protein kinase (SAPK)/c-Jun NH2-terminal kinase (JNK) stress response pathway is stimulated in telomerase-null AEC2 even under normoxic conditions. Expression of downstream apoptotic/stress markers, including caspase-3, caspase-6, Bax, and HSP-25, was also observed in telomerase-null, but not wild-type, AEC2. TUNEL analysis of freshly isolated normoxic AEC2 showed that DNA strand breaks, essentially absent in wild-type cells, increased with each successive terc−/− generation and correlated strongly with telomere length ( R2 = 0.9631). Thus lung alveolar integrity, particularly in the distal epithelial compartment, depends on proper telomere maintenance.

scholarly journals Essential roles for Pot1b in HSC self-renewal and survival

Blood ◽  
2011 ◽  
Vol 118 (23) ◽  
pp. 6068-6077 ◽  
Author(s):  
Yang Wang ◽  
Mei-Feng Shen ◽  
Sandy Chang
Keyword(s):  
Progenitor Cell ◽  
Essential Role ◽  
Dyskeratosis Congenita ◽  
Telomere Maintenance ◽  
Wild Type ◽  
Telomere Shortening ◽  
Damage Response ◽  
First Time

Abstract Maintenance of mammalian telomeres requires both the enzyme telomerase and shelterin, which protect telomeres from inappropriately activating DNA damage response checkpoints. Dyskeratosis congenita is an inherited BM failure syndrome disorder because of defects in telomere maintenance. We have previously shown that deletion of the shelterin component Pot1b in the setting of telomerase haploinsufficiency results in rapid telomere shortening and fatal BM failure in mice, eliciting phenotypes that strongly resemble human syskeratosis congenita. However, it was unclear why BM failure occurred in the setting of Pot1b deletion. In this study, we show that Pot1b plays an essential role in HSC survival. Deletion of Pot1b results in increased apoptosis, leading to severe depletion of the HSC reserve. BM from Pot1bΔ/Δ mice cannot compete with BM from wild-type mice to provide multilineage reconstitution, indicating that there is an intrinsic requirement for Pot1b the maintenance of HSC function in vivo. Elimination of the p53-dependent apoptotic function increased HSC survival and significantly extended the lifespan of Pot1b-null mice deficient in telomerase function. Our results document for the first time the essential role of a component of the shelterin complex in the maintenance of HSC and progenitor cell survival.

Unbalanced X Chromosome Inactivation Independent of Telomere Shortening in Mice Heterozygous for a Mutant Dyskerin Allele.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 184-184
Author(s):  
Baiwei Gu ◽  
Jun He ◽  
Monica Bessler ◽  
Philip J. Mason
Keyword(s):  
Bone Marrow ◽  
Ribosome Biogenesis ◽  
Es Cells ◽  
X Inactivation ◽  
Telomere Maintenance ◽  
Male Mice ◽  
Wild Type ◽  
Telomere Shortening ◽  
Ribonucleoprotein Complexes ◽  
The Impact

Abstract X-linked Dyskeratosis Congenita (DC) is a rare recessive disorder caused by mutations in the DKC1 gene that encodes dyskerin. Dyskerin is part of ribonucleoprotein complexes that participate in two different pathways: ribosome biogenesis and telomere maintenance. It is the subject of intense debate whether disease manifestations in DC are due to dysfunctional telomere maintenances or are caused by a defect in ribosome biogenesis. Pathogenic mutations in dyskerin cause telomere shortening and patients with X-linked DC have critically short telomeres, However, whether there is an additional defect in ribosome biogenesis is difficult to investigate. To dissect the impact of a pathogenic dyskerin mutation on telomeres from the possible additional impact on ribosome biogenesis in an in vivo model, we generated mice expressing a mutant dyskerin protein. Because laboratory mice have very long telomeres a short telomere phenotype requires several generations of inbreeding, whereas a phenotype seen in the first generation is likely to be caused by the defect in ribosome biogenesis. To delete the last 21 amino acids of dyskerin (Del15) we used homologous recombination followed by conditional gene deletion in murine embryonic stem (ES) cells and in mice. Six independent ES cell clones with the deleted Dkc1 gene were obtained. In vitro analysis of the ES cells showed that the Del15 mutation led to dramatically decreased expression of a truncated dyskerin protein with decreased accumulation of the telomerase RNA. In addition, both reduction in telomerase activity and significant telomere shortening after 65 passages were observed. These findings indicate that the Del15 mutation impairs the telomerase maintenance pathway. After testing the accumulation of a series of mouse H/ACA snoRNA in Del15 ES cells, we found a decrease of the mU68 and mE1 snoRNAs suggesting the mutation may also confer effects which are outside the telomerase pathway. We therefore went on to produce a line of mice expressing the truncated Dkc1 protein and were able to obtain male mice hemizygous for the mutant Dkc1 gene as well as female heterozgotes. The male mice express the truncated dyskerin protein and show no gross abnormality up to 6 months of age. Interestingly, heterozygous female mice were healthy as well but the truncated dyskerin protein was dramatically decreased in expression compared to the wild type dyskerin in spleen, thymus, and bone marrow, but not in liver and brain. This result must derive from preferential proliferation of cells expressing wild type dyskerin after random X-inactivation in early embryogenesis. Our analysis indicates that the mutant dyskerin impairs the proliferation in hematopoietic tissues while it does not affect cells which are not rapidly proliferating such as those in liver and brain. Because of the early appearance of the skewed X-inactivation phenotype we conclude that skewing in these mice is caused by a telomere independent mechanism. Interestingly, the lack of overt DC-like abnormalities in the male hemizygous mice indicates that this proliferative disadvantage is insufficient to cause bone marrow failure but in combination with impaired telomere maintenance may accelerate the onset and severity of disease and thus explain the earlier and more severe manifestation in X-linked DC compared to autosomal dominant DC which only affects the telomerase pathway.

TIN2 Mutations In Dyskeratosis Congenita Cause Telomere Shortening In Induced Pluripotent Stem Cells Through Potent Dominant Negative Inhibition Of Telomerase

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 590-590
Author(s):  
Luis Batista ◽  
Franklin Zhong ◽  
Sharon A Savage ◽  
Steven Artandi
Keyword(s):  
Cancer Cells ◽  
Bone Marrow Failure ◽  
Ips Cells ◽  
Dyskeratosis Congenita ◽  
Telomere Maintenance ◽  
Proper Function ◽  
Cell Passage ◽  
Wild Type ◽  
Telomere Shortening ◽  
Telomere Elongation

Abstract Dyskeratosis congenita (DC) is a bone marrow failure syndrome characterized by widespread defects in diverse tissues and a strong predisposition to cancer. DC is caused by germline mutations in genes controlling maintenance of telomeres, nucleoprotein caps that protect chromosome ends. Mutations in components of the telomerase enzyme comprise a large share of cases, including in TERT, TERC, dyskerin, TCAB1, NOP10 and NHP2. These mutations compromise telomerase function leading to telomere shortening, which in turn impairs stem cell function. We previously created patient-derived iPS cells from patients with mutations in TERT, dyskerin or TCAB1 and analyzed these cells to understand the biochemical defects in the telomerase pathway. In each case we found a unique mechanism underlying these telomerase defects, including: reduced catalytic function (TERT mutations), impaired telomerase assembly (dyskerin mutations) and mislocalization of the enzyme to nucleoli (TCAB1 mutations). A six-member protein complex – shelterin - is essential for proper function of telomeres. Despite the critical importance of shelterin proteins in telomere regulation, only a single telomere binding protein – TIN2 – is mutated in DC. However, how these mutations compromise telomere maintenance remains poorly understood. TIN2 mutations occur in a common, autosomal dominant form of DC, presenting in early life, with particularly severe clinical manifestations and poor outcomes. Mutations in the TIN2 gene are clustered in exon 6a, which corresponds to a protein domain of unknown function. To understand how TIN2 mutations impair telomere maintenance and cause DC, we reprogrammed fibroblasts from patients with TIN2 mutations to iPS cells. We succeeded in generating pluripotent iPS cells from a patient with a frame shift mutation at position 284 of the protein. TIN2-mutant iPS cells expressed all the markers of wild-type iPS cells and human ES cells and could be differentiated to all three germ cell layers in culture. With reprogramming from fibroblasts to iPS cells, telomerase is upregulated and causes telomere elongation in wild-type cells. In analyzing telomeres from TIN2-mutant iPS cells, we found that telomere elongation was abrogated. Instead of telomere elongation, TIN2-mutant iPS cells showed telomere shortening with reprogramming and during passage in cell culture. After extended cell passage, TIN2-mutant iPS cells lost the ability to self-renew and differentiated, concomitant with the activation of the telomere surveillance checkpoint p53. To better understand how TIN2 mutant proteins interfere with telomere maintenance, we overexpressed GFP, wild-type TIN2, or TIN2 truncation mutants from DC patients into human, telomerase-positive cancer cells. Genomic DNA was collected from these cells during passage and analyzed for telomere lengths by Southern blot. Expression of GFP or wild-type TIN2 had no effect on telomere lengths, which were stably maintained during the experiment. In marked contrast, expression of the TIN2 truncation mutants from DC patients led to progressive and dramatic telomere shortening with cell passage. Together, these data in patient-derived iPS cells and in human cancer cells suggest that TIN2 mutants inhibit the action of telomerase at telomeres. These results constitute a new molecular mechanism at play in DC and yield new insight into one of the most common forms of DC. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Telomere Length Dynamics and Chromosomal Instability in Cells Derived from Telomerase Null Mice

1999 ◽  
Vol 144 (4) ◽  
pp. 589-601 ◽  
Author(s):  
M. Prakash Hande ◽  
Enrique Samper ◽  
Peter Lansdorp ◽  
María A. Blasco
Keyword(s):  
Telomere Length ◽  
Chromosomal Instability ◽  
Telomere Maintenance ◽  
Chromosome Stability ◽  
Chromosome 2 ◽  
Wild Type ◽  
Telomere Shortening ◽  
Potent Inducer ◽  
Quantitative Fluorescence

To study the effect of continued telomere shortening on chromosome stability, we have analyzed the telomere length of two individual chromosomes (chromosomes 2 and 11) in fibroblasts derived from wild-type mice and from mice lacking the mouse telomerase RNA (mTER) gene using quantitative fluorescence in situ hybridization. Telomere length at both chromosomes decreased with increasing generations of mTER−/− mice. At the 6th mouse generation, this telomere shortening resulted in significantly shorter chromosome 2 telomeres than the average telomere length of all chromosomes. Interestingly, the most frequent fusions found in mTER−/− cells were homologous fusions involving chromosome 2. Immortal cultures derived from the primary mTER−/− cells showed a dramatic accumulation of fusions and translocations, revealing that continued growth in the absence of telomerase is a potent inducer of chromosomal instability. Chromosomes 2 and 11 were frequently involved in these abnormalities suggesting that, in the absence of telomerase, chromosomal instability is determined in part by chromosome-specific telomere length. At various points during the growth of the immortal mTER−/− cells, telomere length was stabilized in a chromosome-specific man-ner. This telomere-maintenance in the absence of telomerase could provide the basis for the ability of mTER−/− cells to grow indefinitely and form tumors.

scholarly journals EXO1 Contributes to Telomere Maintenance in Both Telomerase-Proficient and Telomerase-Deficient Saccharomyces cerevisiae

Genetics ◽  
2004 ◽  
Vol 166 (4) ◽  
pp. 1651-1659 ◽  
Author(s):  
Alison A Bertuch ◽  
Victoria Lundblad
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Proliferation ◽  
Cell Viability ◽  
Telomere Length ◽  
Budding Yeast ◽  
Telomere Maintenance ◽  
Wild Type ◽  
Rapid Loss ◽  
Telomere Shortening ◽  
Telomere Function

Abstract Previous work in budding yeast has indicated that telomeres are protected, at least in part, from the action of Exo1, which degrades the C-rich strand of partially uncapped telomeres. To explore this further, we examined the consequences of Exo1-mediated activity in strains that lacked Ku, telomerase, or both. Loss of Exo1 partially rescued the telomere length defect in a yku80Δ strain, demonstrating that exonuclease action can directly contribute to telomere shortening. The rapid loss of inviability displayed by a yku80Δ est2Δ strain was also partially alleviated by an exo1Δ mutation, further supporting the proposal that Exo1 is one target of the activities that normally protect wild-type telomeres. Conversely, however, Exo1 activity was also capable of enhancing telomere function and consequently cell proliferation, by contributing to a telomerase-independent pathway for telomere maintenance. The recovery of recombination-dependent survivors that arose in a yku80Δ est2Δ strain was partially dependent on Exo1 activity. Furthermore, the types of recombination events that facilitate telomerase-independent survival were influenced by Exo1 activity, in both est2Δ and yku80Δ est2Δ strains. These data demonstrate that Exo1 can make either positive or negative contributions to telomere function and cell viability, depending on whether telomerase or recombination is utilized to maintain telomere function.

scholarly journals Telomerase treatment prevents lung profibrotic pathologies associated with physiological aging

2020 ◽  
Vol 219 (10) ◽  
Author(s):  
Sergio Piñeiro-Hermida ◽  
Chiara Autilio ◽  
Paula Martínez ◽  
Fátima Bosch ◽  
Jesús Pérez-Gil ◽  
...  
Keyword(s):  
Telomere Maintenance ◽  
Alveolar Type ◽  
Proliferative Potential ◽  
Type Ii ◽  
Wild Type ◽  
Telomere Shortening ◽  
Physiological Aging ◽  
Club Cells ◽  
Sporadic Cases ◽  
Deficient Mice

Short/dysfunctional telomeres are at the origin of idiopathic pulmonary fibrosis (IPF) in patients mutant for telomere maintenance genes. However, it remains unknown whether physiological aging leads to short telomeres in the lung, thus leading to IPF with aging. Here, we find that physiological aging in wild-type mice leads to telomere shortening and a reduced proliferative potential of alveolar type II cells and club cells, increased cellular senescence and DNA damage, increased fibroblast activation and collagen deposits, and impaired lung biophysics, suggestive of a fibrosis-like pathology. Treatment of both wild-type and telomerase-deficient mice with telomerase gene therapy prevented the onset of lung profibrotic pathologies. These findings suggest that short telomeres associated with physiological aging are at the origin of IPF and that a potential treatment for IPF based on telomerase activation would be of interest not only for patients with telomerase mutations but also for sporadic cases of IPF associated with physiological aging.

Regulation And Effect Of Telomerase And Telomeric Length In Stem Cells

2020 ◽  
Vol 15 ◽  
Author(s):  
Basak Celtikci ◽  
Gulnihal Kulaksiz Erkmen ◽  
Zeliha Gunnur Dikmen
Keyword(s):  
Telomere Length ◽  
Somatic Cells ◽  
Telomere Maintenance ◽  
The Other ◽  
Telomerase Reverse Transcriptase ◽  
Human Telomerase Reverse Transcriptase ◽  
Telomere Shortening ◽  
Maintenance Mechanism ◽  
Associated Diseases ◽  
Human Telomerase

: Telomeres are the protective end caps of eukaryotic chromosomes and they decide the proliferative lifespan of somatic cells, as the guardians of the cell replication. Telomere length in leucocytes reflects telomere length in other somatic cells. Leucocyte telomere length can be a biomarker of human ageing. The risk of diseases, which are associated with reduced cell proliferation and tissue degeneration, including aging or aging-associated diseases, such as dyskeratosis congenita, cardiovascular diseases, pulmonary fibrosis and aplastic anemia, are correlated with an increase in short telomeres. On the other hand, the risk of diseases, which are associated with increased proliferative growth, including major cancers, is correlated with long telomeres. In most of the cancers, a telomere maintenance mechanism during DNA replication is essential. The reactivation of the functional ribonucleoprotein holoenzyme complex [telomerase] starts the cascade from normal and premalignant somatic cells to advanced malignant cells. Telomerase is overexpressed during the development of cancer and embryonic stem cells, through controlling genome integrity, cancer formation and stemness. Cancer cells have mechanisms to maintain telomeres to avoid initiation of cellular senescence or apoptosis, and halting cell division by critically short telomeres. Modulation of the human telomerase reverse transcriptase is the ratelimiting step for the production of functional telomerase and the telomere maintenance. Human telomerase reverse transcriptase promoter promotes its gene expression only in tumor cells, but not in normal cells. Some cancers activate an alternative lengthening of telomeres maintenance mechanism via DNA recombination to unshorten their telomeres. Not only heritability but also oxidative stress, inflammation, environmental factors, and therapeutic interventions have an effect on telomere shortening, explaining the variability in telomere length across individuals. There have been a large number of publications, which correlate human diseases with progressive telomere shortening. Telomere length of an individual at birth is also important to follow up telomere shortening, and it can be used as biomarkers for healthy aging. On the other hand, understanding of cellular stress factors, which affect stem cell behavior, will be useful in regeneration or treatment in cancer and age-associated diseases. In this review, we will understand the connection between stem cell and telomere biology, cancer, and aging-associated diseases. This connection may be useful for discovering novel drug targets and improve outcomes for patients having cancer and aging-associated diseases.

scholarly journals The History and Mystery of Alveolar Epithelial Type II Cells: Focus on Their Physiologic and Pathologic Role in Lung

2021 ◽  
Vol 22 (5) ◽  
pp. 2566 ◽  
Author(s):  
Barbara Ruaro ◽  
Francesco Salton ◽  
Luca Braga ◽  
Barbara Wade ◽  
Paola Confalonieri ◽  
...  
Keyword(s):  
Host Defense ◽  
Defense Mechanisms ◽  
Work Of Breathing ◽  
Surfactant Proteins ◽  
Alveolar Type ◽  
Type I ◽  
Type Ii ◽  
Lung Protection ◽  
Alveolar Epithelial ◽  
Distal Lung

Alveolar type II (ATII) cells are a key structure of the distal lung epithelium, where they exert their innate immune response and serve as progenitors of alveolar type I (ATI) cells, contributing to alveolar epithelial repair and regeneration. In the healthy lung, ATII cells coordinate the host defense mechanisms, not only generating a restrictive alveolar epithelial barrier, but also orchestrating host defense mechanisms and secreting surfactant proteins, which are important in lung protection against pathogen exposure. Moreover, surfactant proteins help to maintain homeostasis in the distal lung and reduce surface tension at the pulmonary air–liquid interface, thereby preventing atelectasis and reducing the work of breathing. ATII cells may also contribute to the fibroproliferative reaction by secreting growth factors and proinflammatory molecules after damage. Indeed, various acute and chronic diseases are associated with intensive inflammation. These include oedema, acute respiratory distress syndrome, fibrosis and numerous interstitial lung diseases, and are characterized by hyperplastic ATII cells which are considered an essential part of the epithelialization process and, consequently, wound healing. The aim of this review is that of revising the physiologic and pathologic role ATII cells play in pulmonary diseases, as, despite what has been learnt in the last few decades of research, the origin, phenotypic regulation and crosstalk of these cells still remain, in part, a mystery.

The Impact of Age and Gender on Hematopoietic Stem Cells and Immune Contexture of the Bone Marrow Microenvironment

2020 ◽  
pp. 1-6
Author(s):  
Rebar N. Mohammed
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Immune Cells ◽  
Absolute Number ◽  
Cell Number ◽  
Hematopoietic Stem ◽  
The Absolute ◽  
And Gender ◽  
The Impact

Hematopoietic stem cells (HSCs) are a rare population of cells that reside mainly in the bone marrow and are capable of generating and fulfilling the entire hematopoietic system upon differentiation. Thirty-six healthy donors, attending the HSCT center to donate their bone marrow, were categorized according to their age into child (0–12 years), adolescence (13–18 years), and adult (19–59 years) groups, and gender into male and female groups. Then, the absolute number of HSCs and mature immune cells in their harvested bone marrow was investigated. Here, we report that the absolute cell number can vary considerably based on the age of the healthy donor, and the number of both HSCs and immune cells declines with advancing age. The gender of the donor (male or female) did not have any impact on the number of the HSCs and immune cells in the bone marrow. In conclusion, since the number of HSCs plays a pivotal role in the clinical outcome of allogeneic HSC transplantations, identifying a younger donor regardless the gender is critical.

KGF facilitates repair of radiation-induced DNA damage in alveolar epithelial cells

1997 ◽  
Vol 272 (6) ◽  
pp. L1174-L1180 ◽  
Author(s):  
M. Takeoka ◽  
W. F. Ward ◽  
H. Pollack ◽  
D. W. Kamp ◽  
R. J. Panos
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Epithelial Cells ◽  
Dna Polymerase ◽  
Dna Polymerases ◽  
Dna Strand Breaks ◽  
Strand Breaks ◽  
Alveolar Epithelial ◽  
Radiation Induced ◽  
Dna Strand

Administration of exogenous keratinocyte growth factor (KGF) prevents or attenuates several forms of oxidant-mediated lung injury. Because DNA damage in epithelial cells is a component of radiation pneumotoxicity, we determined whether KGF ameliorated DNA strand breaks in irradiated A549 cells. Cells were exposed to 137Cs gamma rays, and DNA damage was measured by alkaline unwinding and ethidium bromide fluorescence after a 30-min recovery period. Radiation induced a dose-dependent increase in DNA strand breaks. The percentage of double-stranded DNA after exposure to 30 Gy increased from 44.6 +/- 3.5% in untreated control cells to 61.6 +/- 5.0% in cells cultured with 100 ng/ml KGF for 24 h (P < 0.05). No reduction in DNA damage occurred when the cells were cultured with KGF but maintained at 0 degree C during and after irradiation. The sparing effect of KGF on radiation-induced DNA damage was blocked by aphidicolin, an inhibitor of DNA polymerases-alpha, -delta, and -epsilon and by butylphenyl dGTP, which blocks DNA polymerase-alpha strongly and polymerases-delta and -epsilon less effectively. However, dideoxythymidine triphosphate, a specific inhibitor of DNA polymerase-beta, did not abrogate the KGF effect. Thus KGF increases DNA repair capacity in irradiated pulmonary epithelial cells, an effect mediated at least in part by DNA polymerases-alpha, -delta, and -epsilon. Enhancement of DNA repair capability after cell damage may be one mechanism by which KGF is able to ameliorate oxidant-mediated alveolar epithelial injury.

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