Interplay between telomere length and telomerase in human leukocyte differentiation and aging.

Blood leukocytes derive from bone marrow hematopoietic stem cells and differentiate into multiple types of mature cells that include granulocytes, monocytes, mast cells of myeloid lineage, and T and B lymphocytes of lymphoid lineage. Their distinctive paths of differentiation and unique roles in immune response provide a model for comparative analysis of biological parameters, such as telomere length and telomerase activity, in different types of leukocytes. Age has also been associated with the decline in immune functions and with the attrition of telomere length in leukocytes. This review will summarize recent progress in the study of telomere length and telomerase expression in leukocytes during differentiation and aging. In addition, I will attempt to shed new light on the roles of telomere and telomerase in leukocyte function and potential clinical interventions.     

Abnormalities in lymphocyte telomerase activity and telomere length in systemic lupus erythematosus]

T-cell telomerase activity was high in the active and inactive stages of systemic lupus erythematosus (SLE). In contrast, B-cell telomerase activity was very high only in the active stage. Compared with normal subjects, SLE patients had a shorter T-cell telomere, but their B-cell telomere length did not differ from that of normal subjects. These findings suggest that T cells are always activated, and that the manifestation of the disease requires the activation of not only T but also B cells. B-cell inhibition alone may be sufficient to suppress the clinical symptoms of SLE, but we consider that the essential treatment of SLE should target T cells as well. In recent years, various biologicals have begun to be used for the treatment of SLE. It is interesting how the use of such biologicals in the future will change T- and B-cell telomerase activity. In formulating a therapeutic strategy using biologicals for SLE, the measurement of telomerase activity in T and B cells seems useful for the preparation of target cells, selection of therapeutic drugs, and evaluation of therapeutic responses.     

Immortal and mortal clonal lymphocyte lines from channel catfish: comparison of telomere length, telomerase activity, tumor suppressor and heat shock protein expression.

Channel catfish autonomous (immortal) and nonautonomous (mortal) leukocyte lines were phenotyped with respect to telomere length and the expression of telomerase, Hsp70 and p53, potentially important factors in cellular immortalization. The autonomous cells constitutively expressed telomerase whereas the nonautonomous cells expressed this activity only transiently. This observation, coupled with the low telomerase activity level seen in freshly isolated leukocytes, suggests that telomerase expression in catfish leukocytes is activation induced. In contrast both types of cell lines exhibited quite similar patterns of significantly shortened telomeres, suggesting that telomerase does not stabilize catfish telomeres until a critical short length is reached. Northern analyses indicated that, like telomerase, Hsp70 gene expression was constitutive in autonomous cells and transient in nonautonomous cells. In contrast, p53 mRNA levels appeared similarly low and noncycling in both long-term cultured types of catfish cells, regardless of the culture situation. Furthermore it was noted, by Western analyses, that both types of cells display multiple sized forms of p53 proteins. This latter observation implies that truncation of p53 protein is probably not directly involved in the in vitro immortalization process of channel catfish leukocytes.     

Lack of correlation between telomere length and telomerase activity and expression in leukemic cells

The expression of three components of telomerase complex (hTR, hTERT, TP1) along with telomerase activity and telomere length in leukemic cells was investigated. Cells were isolated from peripheral blood and/or bone marrow of children with acute lymphoblastic (ALL) and non-lymphoblastic (ANLL) leukemia. Expression of three components of telomerase as well as telomerase activity was found in all leukemic cells. Chemiluminescent detection of terminal restriction fragments (TRF) from DNA isolated from ALL cells showed variable patterns expressing considerable heterogeneity of telomere length. The ALL cells appeared to have both long and short telomere lengths, in contrast to normal peripheral lymphocytes, which produced limited pattern of TRF. The ANLL cells produced predominantly short telomere pattern despite high telomerase activity and expression. It can be concluded that high telomerase activity and expression in leukemic cells is not always correlated with long telomeres (TRF pattern).     

Analyses and comparisons of telomerase activity and telomere length in human T and B cells: Insights for epidemiology of telomere maintenance

Telomeres are the DNA–protein complexes that protect the ends of eukaryotic chromosomes. The cellular enzyme telomerase counteracts telomere shortening by adding telomeric DNA. A growing body of literature links shorter telomere length and lower telomerase activity with various age-related diseases and earlier mortality. Thus, leukocyte telomere length (LTL) and telomerase activity are emerging both as biomarkers and contributing factors for age-related diseases. However, no clinical study has directly examined telomerase activity and telomere length in different lymphocyte subtypes isolated from the same donors, which could offer insight into the summary measure of leukocyte telomere maintenance.We report the first quantitative data in humans examining both levels of telomerase activity and telomere length in four lymphocyte subpopulations from the same donors—CD4+, CD8+CD28+ and CD8+CD28? T cells and B cells, as well as total PBMCs—in a cohort of healthy women. We found that B cells had the highest telomerase activity and longest telomere length; CD4+ T cells had slightly higher telomerase activity than CD8+CD28+ T cells, and similar telomere length. Consistent with earlier reports that CD8+CD28? T cells are replicatively senescent cells, they had the lowest telomerase activity and shortest telomere length. In addition, a higher percentage of CD8+CD28? T cells correlated with shorter total PBMC TL (r = ? 0.26, p = 0.05). Interestingly, telomerase activities of CD4+ and CD8+CD28+ T cells from the same individual were strongly correlated (r = 0.55, r < 0.001), indicating possible common mechanisms for telomerase activity regulation in these two cell subtypes. These data will facilitate the understanding of leukocyte aging and its relationship to human health.     

Telomere length and telomerase activity during expansion and differentiation of human mesenchymal stem cells and chondrocytes

Chondrocyte ex vivo expansion currently performed to replace damaged articular surfaces is associated with a loss of telomeric repeats similar to decades of aging in vivo. This might affect the incidence or time of onset of age-related disorders within transplanted cells or tissues. This study examined whether more immature progenitor cells, such as mesenchymal stem cells (MSC), which can be expanded and subsequently differentiated into chondrocytes is advantageous regarding telomere-length related limitations of expansion protocols. Primary chondrocytes and bone-marrow-derived MSC were isolated from 12 donors, expanded separately to 4 x 10(6) cells, and (re-)differentiated as three-dimensional chondrogenic spheroids. Cells were collected during expansion, after three-dimensional culturing and chondrogenic differentiation, and sequential analyses of telomere length and telomerase activity were performed. Surprisingly, telomeres of expanded MSC were significantly shorter than those from expanded chondrocytes from the same donor (11.4+/-2.5 vs. 13.4+/-2.2 kb) and tended to remain shorter after differentiation in chondrogenic spheroids (11.9+/-1.8 vs. 13.0+/- kb). While telomere lengths in native chondrocytes and MSC were not related to the age of the donor, significant negative correlations with age were observed in expanded (136 bp/year), three-dimensionally reconstituted (188 bp/year), and redifferentiated (229 bp/year) chondrocytes. Low levels of telomerase activity were found in MSC and chondrocytes during expansion and after (re-)differentiation to chondrogenic spheroids. In terms of replicative potential, as determined by telomere length, ex vivo expansion followed by chondrogenic differentiation of MSC did not provide a benefit compared to the expansion of adult chondrocytes. However, accelerated telomere shortening with age during expansion and redifferentiation argues for an "age phenotype" in chondrocytes as opposed to MSC and suggests an advantage for the use of MSC especially in older individuals and protocols requiring extensive expansion     

SENP3对大鼠成骨细胞端粒酶活性及端粒长度的影响

目的:研究sentrin特异性蛋白酶3(SENP3)对大鼠成骨细胞端粒酶活性及端粒长度的影响。方法:首先0.2 mmol/L H_2O_2处理体外培养的大鼠成骨细胞后,Western blotting法检测SENP3及特异性蛋白1(Sp1)的表达。pc DNA3.0-SENP3转染成骨细胞,分别于24 h、48 h、72 h后采用四甲基偶氮唑蓝(MTT)法检测细胞活力的变化。转染48 h后,Western blotting法检测Sp1和端粒酶逆转录酶(TERT)的表达,PCR-TRAP法及PCR法检测端粒酶活性及端粒长度;ELISA检测上清中碱性磷酸酶(ALP)和骨桥蛋白(OPN)的含量;放射免疫法(RIA)检测骨钙蛋白(OCN)的含量。最后将pc DNA3.0-SENP3与siRNA-Sp1共转染成骨细胞,并检测以上指标。结果:H_2O_2处理成骨细胞后,SENP3和Sp1的表达显著上升。pc DNA3.0-SENP3转染成骨细胞后,Sp1和TERT的表达显著上升,细胞活力、ALP、OPN及OCN含量也都显著上升;端粒酶活性显著增加及端粒长度缩短显著延缓。而当pc DNA3.0-SENP3与siRNA-Sp1共转染成骨细胞后,细胞活力,ALP、OPN及OCN含量,端粒酶活性及端粒长度均未发生显著变化。结论:SENP1通过上调Sp1的表达促进TERT的表达,增加端粒酶活性上升及延缓端粒长度缩短,从而增强成骨细胞增殖能力。     

Cell death during lymphocyte development and activation

The development and homeostasis of the immune system requires an exquisite balance between cell proliferation and cell death. In this review, we discuss several in vivo and in vitro models that have been developed to help understand the importance of apoptosis during B and T cell development and activation.     

MiR-138对MCF-7细胞端粒酶活性及端粒稳定性的调控作用

 目的 研究MiR-138通过HTERT作为下游靶基因,对人乳腺癌MCF-7细胞端粒酶活性的调控作用及端粒稳定性的影响。方法 在人乳腺癌MCF-7细胞中瞬时转染MiR-138 模拟物,用MTT法检测细胞增殖活性,并于转染后48h,用实时定量RT-PCR检测端粒酶催化亚单位HTERT表达、TRAP Assay检测端粒酶活性,同时对细胞进行53BP1 抗体免疫荧光染色及端粒的FISH染色。结果 转染后48h,MiR-138模拟物处理的MCF-7细胞HTERT表达水平比对照细胞降低2.18倍(2-△△Ct),端粒酶活性比对照细胞降低2.69倍,53BP1聚集形成的凝集点（Foci）,部分与端粒位点重合,比率达到20.62%±1.55% 。结论 MiR-138以HTERT作为下游靶分子,调控MCF-7细胞端粒酶活性,影响细胞端粒稳定性。     

Attachment and telomere length: more evidence for psychobiological connections between close relationships,health, and aging

Individuals with a history of poor interpersonal relationships are more likely to demonstrate negative health outcomes than those who have had high quality relationships. We sought to evaluate how attachment orientations, stress-induced respiratory sinus arrhythmia (RSA), and self-reported stress were associated with length of telomeres measured from peripheral blood mononuclear cells. Participants (N = 213) completed self-report measures of attachment and stress. Measurement of RSA was conducted before and after a stressful task and a blood draw was completed for analysis of telomere length. Attachment orientations were not directly associated with telomere length; however, we found that high attachment anxiety was associated with shorter length of telomeres via high self-reported stress. Attachment avoidance was also associated with telomere length via self-reported stress, but only among those with high stress-induced RSA. Exploratory analyses of T cell subsets indicated that stress was most strongly associated with telomeres from CD8CD28+ cells in comparison to CD8CD28? and CD4 cells. Study findings indicate that attachment orientations are associated with telomere length via stress, providing novel insights into the mechanisms through which close relationships can impact health and aging.     

A tale of two TRAPs: LAT and LAB in the regulation of lymphocyte development, activation, and autoimmunity

Transmembrane adaptor proteins (TRAPs) link antigen receptor engagement to downstream cellular processes. Although these proteins typically lack intrinsic enzymatic activity, they are phosphorylated on multiple tyrosine residues following lymphocyte activation, allowing them to function as scaffolds for the assembly of multi-molecular signaling complexes. Among the many TRAPs that have been discovered in recent years, the LAT (linker for activation of T cells) family of adaptor proteins plays an important role in the positive and negative regulation of lymphocyte maturation, activation, and differentiation. Of the two members in this family, LAT is an indispensable component controlling T cell and mast cell activation and function; LAB (linker for activation of B cells), also called NTAL, is necessary to fine-tune lymphocyte activation and may be a key regulator of innate immune responses. Here, we review recent advances on the function of LAT and LAB in the regulation of development and activation of immune cells.     

Developmental regulation of telomerase activity in human fetal tissues during gestation

Telomerase is a ribonucleoprotein that adds hexanucleotide repeats (telomeres) to the ends of linear chromosomes, compensating for the loss of telomeric DNA which occurs with DNA replication. In humans, telomerase has been previously detected in germ-line tissues, blastocysts, 16-20 week old fetal tissue, and most cancers, but not in mature sperm or ova, or in most normal somatic tissues. It has been hypothesized that telomerase is suppressed during somatic development and reactivated in malignancy. To test the hypothesis that telomerase is suppressed during somatic development, human fetal tissues of 8-21 weeks gestational age were assayed for telomerase activity. All tissues expressed telomerase at the earliest ages examined. Lung, liver, spleen, and testis maintained telomerase activity through the latest age assayed, namely 21 weeks. Brain and kidney telomerase activity was present up to the 16th week and was undetectable thereafter. Heart tissue did not display activity beyond the 12th week. Lysates of heart, brain, and kidney without telomerase activity did not inhibit the activity of known telomerase-positive cells, suggesting that suppression of telomerase activity during gestational development is due to a lack of active telomerase rather than to the presence of an inhibitor. These findings demonstrate tissue-specific and developmental regulation of telomerase in the human fetus, suggesting an important role for this ribonucleoprotein in human fetal tissue differentiation and development.