Thyroglobulin may affect telomerase activity in thyroid follicular cells

Telomerase (TA) activity is known to be present in malignant tumor cells, but not in most somatic differentiated cells. TA shows relatively high activity in thyroid cancer cells, but reports vary. This fact prompted us to elucidate whether cell component inhibitors of TA in the thyroid follicles can modulate its activity. The activity of TA extracted from Hela cells was inhibited by mixing with the supernatant fraction of human thyroid tissue extract. To examine the effect of iodine, thyroid hormones (?-T3 and ?-T4) and human thyroglobulin (hTg) contained in the thyroid follicles, ?-T3, ?-T4 and hTg were added to the TRAP assay system in vitro, using TA from Hela cells. Iodine, ?-T3 and ?-T4 did not affect TA activity, but hTg inhibited the TA activity in a dose-dependent manner (IC₅₀ of hTg: ca 0.45 μM: inhibiting concentration of hTg was from 0.15 μM to 3.0 μM). The hTg inhibition was not evident in the RT-PCR system, suggesting no effect of hTg on Taq DNA polymerase activity. The hTg inhibition of TA activity was attenuated by dNTP but not significantly by TS primer. These data suggest that hTg contained in thyroid follicular cells of various thyroid diseases may affect the TA activity measured in biopsied thyroid specimens, and that the reduction of the TA activity by hTg may induce slow progression and growth, and low grade malignancy of thyroid cancer, particularly differentiated carcinoma.     

Telomerase with mutated catalytic motifs has dominant negative effects on telomerase activity and inhibits cell growth

Telomerase catalytic subunit (TERT) seems a key factor controlling telomerase activity, telomere length, and cell growth. To further address this issue, we forced expression of a catalytically inactive mutant human TERT (hTERT) in hTERT-immortalised sheep fibroblasts to examine its effects. Expression of mutant hTERT compromised telomerase activity reconstituted by wild-type hTERT in a manner directly attributable to mutant hTERT expression level. High levels of mutant hTERT expression inhibited cell growth with a subset of cells entering replicative senescence. Furthermore, significant telomere attrition was evident in two of three clones with high levels of mutant hTERT expression. Our findings are consistent with the notion that hTERT homodimers are necessarily required to form a functional telomerase complex at the telomere substrate. We also highlight the requirement of a more thorough understanding of telomerase- and telomere-associated factors to understand fully the interplay that governs telomere homeostasis in vitro and in vivo.     

P311 binds to the latency associated protein and downregulates the expression of TGF-beta1 and TGF-beta2

P311 is an 8-kDa protein originally found in neurons and muscle. We recently showed that expression of P311 in NIH 3T3 cells induced a myofibroblast phenotype with low TGF-beta1 expression. Here we demonstrate that P311 downregulates not only TGF-beta1, but also TGF-beta2, expression, with no effect on TGF-beta3. In addition, P311 interacts with TGF-beta2 in a yeast two-hybrid system through a sequence encompassing part of the TGF-beta latent associated protein (LAP) and part of mature TGF-beta2. Coimmunoprecipitations demonstrated interaction between P311 and TGF-beta1 and 2, but not TGF-beta3. Additional coimmunoprecipitations after introducing LAP or mature TGF-beta1 into cells demonstrated P311 binding to LAP, but not to mature TGF-beta. P311 has a conserved PEST domain, which generally serves as a rapid degradation signal. Deletion of the PEST domain reversed the effect of P311 on TGF-beta isoforms. Finally, Smad3 activity was decreased in P311-expressing cells, but was corrected by exogenous TGF-beta1 treatment, which also elevated TGF-beta1 mRNA level. This suggested that P311 downregulates TGF-beta1 and 2 in part by blocking TGF-beta autoinduction.     

IGFBP7在甲状腺乳头状癌中的表达及意义

目的观察胰岛素样生长结合蛋白-7(IGFBP7)在甲状腺乳头状癌中的表达及意义。方法对87例甲状腺乳头癌(PTC)组织及癌旁组织(PeT)进行IGFBP7、Ki67、p53免疫组织化学分析,其中10例提取蛋白用半定量western blot方法进行IGFBP7表达水平分析。结果 (1)IGFBP7免疫组化显示,PTC癌组织IGFBP7阳性率达78.16%(68/87),明显高于癌旁组织的31.03%(27/87)(P0.05);(2)Western blot灰度值比较,癌组织IGFBP7表达水平显著高于癌旁组织(t=2.875,P0.05);(3)Ki67、p53与IGFBP7相关性分析显示,p53表达水平与IGFBP7表达水平呈显著正相关性(r=0.261,P0.05),而Ki67表达水平与IGFBP7表达水平无相关性(r=0.148,P=0.170);(4)高水平表达的IGFBP7与PTC淋巴结转移高度相关(r=0.238,P0.05)。结论 IGFBP7表达异常增高可能与甲状腺乳头状癌发生及发展有关,有望成为诊断PTC的一项新型的生物标记物。     

Uridine phosphorylase 1 associates to biological and clinical significance in thyroid carcinoma cell lines

Thyroid cancer incidence has been continuity increasing worldwide. Uridine phosphorylase 1 (UPP1) is a protein‐coding gene and has been detected that UPP1 was the higher expression in many solid malignancies, just as head and neck cancers, breast cancer, compared with paired normal tissue. But the act of UPP1 in thyroid cancer is not explicit. In this article, we investigate the function of UPP1 expression in thyroid cancer. The Cancer Genome Atlas (TCGA) unpaired thyroid cancer and normal RNA‐seq data were downloaded, and our paired thyroid cancer and normal samples were analysed by a polymerase chain reaction. The expression of UPP1 was regulated by transfected small interfering RNA, and the function of UPP1 was determined via migration, invasion and cell proliferation assays. Western blot assay was achieved to determine the UPP1 expression correlates with the function of 5‐FU regulate epithelial‐mesenchymal transition. The significant upregulation of UPP1 in thyroid cancer tissues compared with normal thyroid tissues was revealed by our data and TCGA data. UPP1 overexpression was significantly correlated with lymph node metastasis, tumour stage and tumour size. In the cell, experiments showed that UPP1 low expression significantly suppressed the migration, invasion and proliferation. Western blot assay proves the effect of UPP1 expression on 5‐FU regulates epithelial‐mesenchymal transition pathway. UPP1 plays a crucial oncogene in thyroid cancer. Our findings indicate that UPP1 might be a biomarker of thyroid cancer and may act by regulating epithelial‐mesenchymal transition (EMT).     

The role of telomeres and telomerase in stem cell aging

Stem cells regenerate our bodies. In a similar manner to match ignition, stem cell “ignition” has to be precisely tuned to avoid uncontrolled proliferation as may occur in tumors or, inversely, the lack of proliferation as happens in degenerative disorders. During the last years it has become evident that telomeres and telomerase are main components of the stem cell “ignition” mechanism, providing a way to restrain cancer and delay aging.     

The effects of G-CSF and naproxen sodium on the serum TGF-beta1 level and fracture healing in rat tibias

Local and systemic release of transforming growth factor beta 1 (TGF-beta1) is known to increase during the process of fracture healing and this cytokine stimulates bone healing. The majority of the non steroidal anti inflammatory drugs (NSAIDs) inhibit fracture healing. Granulocyte colony stimulating factor (G-CSF) is a hematopoietic growth factor that stimulates bone marrow. In this study, the effects of the NSAID naproxen sodium, G-CSF, and both of them in combination on the TGF-beta1 serum level in rats with tibia fractures were measured and fracture healing was evaluated by histopathologic and radiologic examination. The TGF-beta1 serum levels obtained on day one (24 h after fracture but before administration of naproxen or G-CSF) were found to be similar in all of the five groups (p > 0.05). At the end of the first week, TGF-beta1 levels were significantly lower in naproxen-treated rats than those of the other groups excluding control (p = 0.002). Similar changes in TGF-beta1 levels were found at the end of the second and fourth weeks. TGF-beta1 levels were significantly higher in G-CSF-treated rats at the end of the first, second and fourth weeks (p < 0.05). Fracture healing scores measured with histopathological and radiological methods were higher in G-CSF-treated rats than in naproxen-treated ones. When both naproxen and G-CSF were given, the scores resumed to normal. The results point to the negative effect of naproxen sodium on fracture healing is due to its decreasing effect on the level of TGF-beta1, which may be a new possible mechanism. Moreover, this negative effect can be inhibited by the use of G-CSF.     

Role of Dicer1 in thyroid cell proliferation and differentiation

DICER1 plays a central role in the biogenesis of microRNAs and it is important for normal development. Altered microRNA expression and DICER1 dysregulation have been described in several types of tumors, including thyroid carcinomas. Recently, our group identified a new somatic mutation (c.5438A>G; E1813G) within DICER1 gene of an unknown function. Herein, we show that DICER1 is overexpressed, at mRNA level, in a significant-relative number of papillary (70%) and anaplastic (42%) thyroid carcinoma samples, whereas is drastically downregulated in all the analyzed human thyroid carcinoma cell lines (TPC-1, BCPAP, FRO and 8505c) in comparison with normal thyroid tissue samples. Conversely, DICER1 is downregulated, at protein level, in PTC in comparison with normal thyroid tissues. Our data also reveals that DICER1 overexpression positively regulates thyroid cell proliferation, whereas its silencing impairs thyroid cell differentiation. The expression of DICER1 gene mutation (c.5438A>G; E1813G) negatively affects the microRNA machinery and cell proliferation as well as upregulates DICER1 protein levels of thyroid cells but has no impact on thyroid differentiation. In conclusion, DICER1 protein is downregulated in papillary thyroid carcinomas and affects thyroid proliferation and differentiation, while DICER1 gene mutation (c.5438A>G; E1813G) compromises the DICER1 wild-type-mediated microRNA processing and cell proliferation.     

Immortalization with telomerase of the Nestin-positive cells of the human pancreas

Cells expressing the neuronal stem cell marker Nestin are present in the human pancreas but the biological role of these cells has yet to be resolved. We report here the establishment with the catalytic subunit of human telomerase (hTERT) of a line of normal human cells representing this cell type. Primary human cells derived from the ducts of the pancreas were transduced with an hTERT cDNA. The infected cells became positive for telomerase, failed to senesce, and were still proliferating after more than 150 doublings. The immortalized cells were positive for the expression of Nestin (at both the mRNA and protein levels) and were found to be free of cancer-associated changes: diploid and expressing wild type p16(INK4a), p53, and K-Ras. An established line of normal human cells representing this cell type should be of great value to help define the biological properties of this novel cell type.     

Differential regulation of osteoadherin (OSAD) by TGF-beta1 and BMP-2

Osteoadherin (OSAD) is a member of the small leucine rich-repeat proteoglycan (SLRP) family. SLRPs are normally found in extracellular matrices, but OSAD is the only member restricted to mineralized tissues. We investigated the promoter region of OSAD by in silico analysis and found that the proximal promoter region contains sites for Smad-3, Smad-4, and AP-1. All are effectors of TGF-beta family signalling. We tested sensitivity of the promoter to the two TGF-beta family members TGF-beta1 and BMP-2. We found TGF-beta1 to down regulate OSAD, while BMP-2 up regulates OSAD. As a consequence of how OSAD is regulated by TGF-beta1 and BMP-2 and its temporal expression pattern in osteoblasts and bone development, we can conclude OSAD as an early marker for terminally differentiated matrix producing osteoblasts.     

Genomics of TGF-beta1 signaling in stem cell commitment and dendritic cell development

Dendritic cells are professional antigen presenting cells and central for establishing and maintaining immunity and immunological tolerance. They develop from hematopoietic stem cells through successive steps of lineage commitment and differentiation. Dendritic cell development and function are regulated by specific cytokines, including transforming growth factor type beta1 (TGF-beta1). Our previous work demonstrated the importance of TGF-beta1 signaling for dendritic cell development and subset specification. Here, we used genome-wide gene expression profiling with DNA microarrays to investigate the activity of TGF-beta1 on gene expression in dendritic cell development. This study identified specific gene categories induced by TGF-beta1 with an impact on dendritic cell biology.     

MicroRNA-524-5p suppresses the progression of papillary thyroid carcinoma cells via targeting on FOXE1 and ITGA3 in cell autophagy and cycling pathways

MicroRNAs are beneficial for cancer therapy as they can simultaneously downregulate multiple targets involved in diverse biological pathways related to tumor development. In papillary thyroid cancer, many microRNAs were identified as differentially expressed factors in tumor tissues. In another way, recent studies revealed cell proliferation, cell cycling, apoptosis, and autophagy are critical pathways controlling papillary thyroid cancer development and progression. As miR-524-5p was approved as a cancer suppressor targeting multiple genes in several types of cancer cells, this study aims to characterize the role of miR-524-5p in the thyroid cancer cell. The expression of miR-524-5p was decreased in the papillary thyroid cancer tissues and cell lines, while forkhead box E1 (FOXE1) and ITGA3 were increased. In the clinical case, expression of miR-524-5p, FOXE1, and ITGA3 were significantly correlated with papillary thyroid cancer development and progression. FOXE1 and ITGA3 were approved as direct targets of miR-524-5p. miR-524-5p could inhibit papillary thyroid cancer cell viability, migration, invasion, and apoptosis through targeting FOXE1 and ITGA3. Cell cycling and autophagy pathways were disturbed by downregulation of FOXE1 and ITGA3, respectively. Collectively, miR-524-5p targeting on FOXE1 and ITGA3 prevents thyroid cancer progression through different pathways including cell cycling and autophagy.