MiR-30a inhibits osteolysis by targeting RunX2 in giant cell tumor of bone

RunX2 has been identified to crucially regulate the osteolysis in giant cell tumor of bone. MiR-30a is an intronic miRNA identified as tumor suppressor, but little is known about its role in giant tumor cell of bone. In our research, we reported miR-30a was down-regulated in GCT whereas RunX2 was highly expressed. Further research proved that miR-30a can regulate the expression of RunX2 by binding to its 3′-UTR, which influence the osteoclast differentiation and osteolysis formation. Thus, these results suggest that miR-30a could directly target RunX2 and participate in osteolysis in GCT.     

Expression and localization of extracellular matrix metalloproteinase inducer in giant cell tumor of bone

Matrix metalloproteinases (MMPs) are regarded as a significant regulator in tumor invasion and metastasis. Previous studies have shown that extracellular matrix metalloproteinase inducer (EMMPRIN) in tumor cells induces the synthesis of MMPs. EMMPRIN is abundantly present on the surface of tumor cells and stimulate adjacent stromal cells to synthesize MMPs to induce tumor progression. Giant cell tumor (GCT) of bone is a benign but locally aggressive primary neoplasm of bone. The spindle-shaped mononuclear stromal cells are considered to be the tumor components of GCT, which are capable of inducing osteoclast formation by recruiting the circulating monocyte and macrophage. In this study, we proposed that EMMPRIN is associated with the biological progression and aggressiveness of GCT. We have conducted semi-quantitative RT-PCR to determine the correlation of EMMPRIN expression with the clinical stage of GCT. We have also examined the cellular localization of EMMPRIN in GCT using in-situ hybridization (ISH) and Immunohistochemistry (IH). The results showed that EMMPRIN was present in GCT and its mRNA levels were associated with the clinical stage of GCT. Higher expression level of EMMPRIN was observed in GCT with advanced stage (stage III). There was a great significance (P < 0.05) of EMMPRIN expression between stage I & II and stage III GCTs. Both ISH and IH demonstrated that EMMPRIN is present at the multinuclear osteoclast-like giant cells of GCT, with strong immunostaining on the cell membrane. The stromal-like tumor cells were also positively stained but the intensity was weaker. Interestingly, the production of EMMPRIN in osteoclast-like cells of GCT seems to be regulated by stromal-like tumor cells. Receptor activator of NF-kappaB ligand (RANKL), which has been previously shown to be produced by the stromal-like tumor cells for the recruitment of osteoclast-like giant cells in GCT, enhanced the expression of EMMPRIN mRNA during the differentiation of macrophage-like RAW(264.7) cells into osteoclasts. In short, our studies suggest that EMMPRIN may be an important regulatory factor involved in the biological behaviors of GCT.     

PP2A as a master regulator of the cell cycle

Protein phosphatase 2A (PP2A) plays a critical multi-faceted role in the regulation of the cell cycle. It is known to dephosphorylate over 300 substrates involved in the cell cycle, regulating almost all major pathways and cell cycle checkpoints. PP2A is involved in such diverse processes by the formation of structurally distinct families of holoenzymes, which are regulated spatially and temporally by specific regulators. Here, we review the involvement of PP2A in the regulation of three cell signaling pathways: wnt, mTOR and MAP kinase, as well as the G1→S transition, DNA synthesis and mitotic initiation. These processes are all crucial for proper cell survival and proliferation and are often deregulated in cancer and other diseases.     

The surface membrane as a regulator of animal cell division

Summary The surface membrane of an animal cell is proposed to be the target for regulation of cell division. It undergoes regular periodic changes during the cell division cycle. Interference with these changes by cell-cell surface contacts is proposed to prevent the normal progression of events, and thereby can change the metabolic pattern so as to put the cells into a resting state. Through external influences, cells can escape from this resting state; when this occurs surface changes are the earliest ones observed. Cells that have become malignant, particularly after virus infection, show marked changes in their surface properties. Infection is proposed to prevent the surface changes that lead to the resting state. Recent evidence from in vitro experiments is summarized, and some speculations are made on the connection between the surface and processes of division such as nuclear replication. Presented in the Symposium on Regulation in Tumor Cells at the 22nd Annual Meeting of the Tissue Culture Association. Lake Placid, New York. This work was supported by Public Health Service Grant CA-A1-1195 and Grant E-555 from the American Cancer Society.     

Establishment of functional human pituitary tumor cell cultures

Summary Five primary human pituitary tumor cell cultures were initiated from adenoma fragments obtained from patients with prolactin-secreting adenomas and acromegaly. Functional cell cultures were maintained and propagated in monolayer or suspension culture for up to 9 months. Optimal cell viability and growth were achieved using Ham’s F10 medium enriched with 20% fetal bovine serum, although cells from a patient with acromegaly also grew in serum-free, defined, hormone-containing medium. Bromocriptine (100 ng/ml) did not alter the growth curve of replicating cells derived from a patient with acromegaly. These cells initially secreted 5.5 μg human growth hormone/10⁶ cells, and hormone production diminished after 6 wk. Prolactin secretion by cells derived from prolactinomas (0.5 to 1.3 μg/10⁶ cells/24 h) was stimulated by thyrotropin-releasing hormone (10 ng/ml) in two of the cultures. Both dopamine (10 ng/ml) and nickel chloride (1 mM) suppressed PRL secretion. These studies demonstrate that responsive human pituitary tumor cell cultures can be initiated and maintained. This research was supported by VA Medical Research Funds and NIH Grant HD 7181.     

Possible role of cell cycle-dependent morphology,geometry, and mechanical properties in tumor cell metastasis

Studies that examine the shear- and abrasion-sensitivity of proliferating cells are important in order to understand the behavior of hybridoma cells in bioreactor culture and metastasizing cancer cells in the bloodstream. Little is known about the link between morphology, structure, and mechanical properties of a given cell line, especially with respect to variations throughout the cell cycle. In our experiments with GAP A3 hybridoma cells, distinct cell morphologies were identified and correlated with phases of the cell cycle by video microscopic observation of synchronized cells, and of individual cells that were followed throughout their cell cycle. Micropipet manipulation was used to measure the geometrical (cell volume) and mechanical (apparent cell viscosity) properties of single cells. As the cell cycle progressed at 37°C, an increase in cell volume from 1400 μm³ to 5700 μm³ was accompanied by an increase in apparent cell viscosity from 430 poise to 12,000 poise, consistent with an accumulation of more cytoplasmic material in the “older” cells. Hybridomas are representative of the various leukemias derived from hemopoietic cells, and even though as a whole, they appeared to be rather shear-insensitive, the wide range of property values demonstrates that a given cell line cannot be characterized by a single value for any one property, and that properties must be related to the cell cycle when considering proliferating cells. It is interesting to see if distinct stages in the metastatic sequence of events might correlate with any of these physical features of the cell cycle, irrespective of cell type or cell line. For example, the cytokinetic doublet could represent a fragile structure that may fail and produce cell death under fluid-shear conditions that would not affect the cells at any other stage in the cell cycle. Identifying such cell cycle-dependent features in metastasizing cancer cells could lead to a better understanding of the metastatic process and to possible clinical treatments directed at making cells more shear- and abrasion-sensitive, and therefore, more likely to be killed by the natural hydrodynamic forces of the circulatory system.     

Activation of AMP kinase plays a role in the increased apoptosis in the renal proximal tubule in cystinosis

In cystinosis, renal proximal tubule (RPT) function is compromised, due to mutations in ctns, which encodes for the transporter cystinosin, which removes cystine from lysosomes. Altered RPT function in cystinosis has been attributed to decreased ATP, as well as increased apoptosis. In this report, the role of AMPK was examined. AMPK was activated in primary rabbit RPT cells with a cystinosin knockdown, using cystinosin siRNA. The activation of AMPK was associated with a 50% decrease in ATP and a 1.7-fold increase in the ADP/ATP level. Cisplatin-induced apoptosis also increased in primary RPT cells with a cystinosin knockdown. The role of AMPK in the increased sensitivity to cisplatin was examined. The increased sensitivity to cisplatin was prevented in primary RPT cells with a cystinosin knockdown by the AMPK inhibitor Compound C. The effect of siRNAs against AMPKα1 and AMPKα2 was also studied. The siRNAs knocked down AMPKα, and prevented AMPKα activation by 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR). The siRNAs against AMPKα1 and AMPKα2 also prevented the increased sensitivity to cisplatin in the primary RPT cells with a cystinosin knockdown. These results suggest that signaling through AMPK plays a role in the enhanced apoptosis in the RPT in cystinosis.     

ThPOK在T细胞分化发育中的作用

ThPOK（T-helper-inducing POZ/Krueppel-like factor）又被称为Zbtb7b、Zfp67、cKrox,隶属于一个很大的转录因子家族——POK家族。ThPOK最初是被认为与Ⅰ型胶原蛋白基因的转录抑制有关,但近年来的研究发现,ThPOK在T细胞分化过程中至关重要,特别是对CD4^＋T细胞的分化发育起着命运决定的核心作用。该文综述了ThPOK在CD4^＋T细胞分化过程中的作用特点及其与另外两种重要转录因子GATA3和Runx3的相互作用关系,并在此基础上阐述了ThPOK在其他T细胞,如iNKT细胞、γδT细胞及效应CD8^＋T细胞中的作用功能。     

The role of the microenvironment in tumor immune surveillance

The evidence appears compelling that the microenvironment, and associated biological cellular and molecular factors, may contribute to the progression of a variety of tumors. The effects of the microenvironment may directly influence the plasticity of T cell lineages, which was recently discussed (O''Shea & Paul, 2010 [4]). To review the putative role of the microenvironment in modulating the commitment of tumor immune surveillance, we use the model of oral premalignant lesions.     

TWIST1在人脐静脉内皮细胞增殖、迁移及血管形成中的作用

探讨TWIST1在原代人脐静脉内皮细胞（human umbilical vein endothelial cells,HUVECs）增殖、迁移及体外血管生成中的作用。用有靶向人TWIST1基因shRNA（pLL3.7-shTwist1-GFP）的慢病毒液感染试验组细胞,同时以携带Scramble shRNA的慢病毒液（pLL3.7-shCtrl-GFP）感染对照组细胞,用流式细胞术测定细胞感染效率,实时荧光定量PCR（real-time fluorescent quantitative PCR,qRT-PCR）检测shRNA的基因沉默效率。通过制作细胞生长曲线、Annexin V/7AAD染色流式细胞术、细胞划痕实验、小管形成实验、qRT-PCR检测TWIST1表达降低对HUVECs的增殖、凋亡、迁移、血管形成能力以及血管生长因子受体2（vascular endothelial growth factor receptor 2,VEGFR2）基因表达的影响。试验组TWIST1基因表达下降为对照组的30%,表明shTWIST1能有效降低TWIST1基因的表达。与对照组相比,敲降TWIST1能明显抑制HUVECs的增殖（P<0.01）,诱导细胞凋亡（P<0.05）。试验组HUVECs划痕愈合率、体外生成的血管样结构数目和总小管分支长度均显著低于对照组（P<0.01）;与对照组相比,试验组HUVECs中VEGFR2的表达显著降低（P<0.01）。通过探究HUVECs表达的TWIST1在内皮细胞增殖、存活、迁移和毛细血管样结构的形成中的作用,为TWIST1作为抑制肿瘤血管新生治疗的新靶点提供一定的理论依据。     

Siglec-15, a member of the sialic acid-binding lectin, is a novel regulator for osteoclast differentiation

Osteoclasts are tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells derived from monocyte/macrophage-lineage precursors and are critically responsible for bone resorption. In giant cell tumor of bone (GCT), numerous TRAP-positive multinucleated giant cells emerge and severe osteolytic bone destruction occurs, implying that the emerged giant cells are biologically similar to osteoclasts. To identify novel genes involved in osteoclastogenesis, we searched genes whose expression pattern was significantly different in GCT from normal and other bone tumor tissues. By screening a human gene expression database, we identified sialic acid-binding immunoglobulin-like lectin 15 (Siglec-15) as one of the genes markedly overexpressed in GCT. The mRNA expression level of Siglec-15 increased in association with osteoclast differentiation in cultures of mouse primary unfractionated bone marrow cells (UBMC), RAW264.7 cells of the mouse macrophage cell line and human osteoclast precursors (OCP). Treatment with polyclonal antibody to mouse Siglec-15 markedly inhibited osteoclast differentiation in primary mouse bone marrow monocyte/macrophage (BMM) cells stimulated with receptor activator of nuclear factor κB ligand (RANKL) or tumor necrosis factor (TNF)-α. The antibody also inhibited osteoclast differentiation in cultures of mouse UBMC and RAW264.7 cells stimulated with active vitamin D₃ and RANKL, respectively. Finally, treatment with polyclonal antibody to human Siglec-15 inhibited RANKL-induced TRAP-positive multinuclear cell formation in a human OCP culture. These results suggest that Siglec-15 plays an important role in osteoclast differentiation.     

In vivo test system for tumor production by cell lines derived from lower vertebrates

Summary The immune suppressed lizard,Anolis carolinensis, can be used to test for in vivo tumor production by cell lines derived from a variety of ectothermic vertebrates. Cell lines tested for tumor production were also assessed for loss of attachment-dependent proliferation and contact inhibition of cell overlap. The results demonstrate that the criteria standardly used to assess transformation and neoplastic change in cultured mammalian cells apply equally well to cultured cells from ectotherms. Supported by grants AG01476 and NS24162 from the National Institutes of Health, Bethesda, MD.     

Progesterone as a regulator of granulosa cell viability

Progesterone (P4) prevents numerous cells, including uterine, mammary and ovarian cells, from undergoing apoptosis. Interestingly, P4 prevents apoptosis of ovarian granulosa cells (GCs), which do not express the classic nuclear P4 receptor. This review presents data that support a non-genomic action of P4 in granulosa cells. These studies were conducted using both primary rat granulosa cells and rat spontaneously immortalized granulosa cells (SIGCs). Specifically, these studies reveal that (1) ³H-P4 specifically binds to SIGCs; (2) an antibody directed against the ligand binding domain of the nuclear P4 receptor (C-262) detects a 60 kDa protein, which localizes to the plasma membrane and binds P4; and (3) treatment with C-262 blocks P4’s ability to maintain granulosa cell viability. Additional studies demonstrate that a protein kinase G (PKG) activator, 8-br-cGMP, mimics and PKG antagonists, Rp-8-pcCPT-GMP and KT5823, attenuate P4’s action. These studies support the concept that the 60 kDa P4 binding protein functions as membrane receptor for P4 which activates a PKG-dependent mechanism to regulate granulosa cell survival.     

肿瘤细胞侵袭研究进展

肿瘤细胞侵袭和转移是癌医学和癌生物学最重要的难题,癌症主要因其肿瘤细胞的侵袭和转移而成为致命的疾病,虽然侵袭和转移的机制仍不清楚,但肿瘤细胞侵袭一直是研究热点,本文就近年来对肿瘤细胞侵袭研究的新进展进行综述,以期为寻找治疗肿瘤的新方案提供参考.