西洋参叶二醇组皂苷抑制血管平滑肌细胞增殖及影响原癌基因表达的研究

观察PQDS对VSMC增殖及原癌基因c-myc的mRNA表达的影响,探讨PQDS抑制VSMC的增殖作用及相关机制。方法用组织贴块法培养大鼠胸主动脉VSMC作为研究对象,用AngⅡ诱导VSMC增殖,随机分为：空白对照组、AngⅡ模型组、AngⅡ＋PQDS 25、50、100mg/L不同剂量组。用MTT法、流式细胞术、RT-PCR法观察PQDS对细胞增殖、细胞周期、增殖指数及原癌基因c-myc的mRNA表达的影响。结果PQDS可减低VSMC的增殖能力,将增殖的VSMC抑制在G0/G1期,可以下调VSMC中的原癌基因c-myc的mRNA表达。结论PQDS能抑制VSMC增殖,其机制可能与下调原癌基因c-myc的表达有关。     

Chromosomal localization of the human proto-oncogene c-ets

E26 is an acute leukaemia avian retrovirus which induces myeloblastosis and erythroblastosis in vivo and transforms erythroblasts and myeloblasts in vitro. It contains the oncogene v-myb (ref. 4), first described for avian myeloblastosis virus (AMV), as well as a second specific nucleotide sequence, v-ets located 3' to v-myb (refs 5,6). We have reported that v-ets has a cellular counterpart (c-ets) in chicken and human DNA. Now, using two independent methods--hybridization with human c-ets probe of sorted chromosomes and in situ hybridization--we report the localization of the ets locus on human chromosome 11 at bands q23-q24. This finding may be important, as specific breakpoints around this position have been reported for human malignancies such as acute monocytic leukaemia and Ewing's sarcoma.     

Requirement for ras proto-oncogene function during serum-stimulated growth of NIH 3T3 cells

Human tumours often contain DNA sequences not found in normal tissues which are able to transform cultured NIH 3T3 cells. In some tumours the gene responsible for this transformation belongs to the cellular ras gene family. A specific type of mutation is responsible for converting the cellular proto-oncogene into a ras oncogene capable of inducing transformation. In a study of the function of a cellular ras gene, its protein product (produced in a bacterial cell) was microinjected into NIH 3T3 cells; the recipient cells became morphologically transformed and were induced to initiate DNA synthesis in the absence of added serum, but only when cellular ras protein was injected at much higher concentrations than required with protein of the transforming ras gene. To further analyse the function of the cellular ras gene, we have now injected monoclonal antibodies against ras proteins into NIH 3T3 cells. We report here that NIH 3T3 cells induced to divide by adding serum to the culture medium are unable to enter the S phase of the cell cycle after microinjection of anti-ras antibody, showing that the protein product of the ras proto-oncogene is required for initiation of the S-phase in NIH 3T3 cells.     

MicroRNA-25对大鼠骨关节炎软骨细胞增殖和凋亡调节作用

探讨miR-25对白细胞介素1β(IL-1β)促进OA软骨细胞凋亡和增殖的影响。以木瓜蛋白酶膝关节腔注射诱导骨性关节炎动物模型和SD大鼠骨性关节炎软骨细胞原代培养。IL-1β体外刺激OA软骨细胞,模拟骨性关节炎体外环境。MiR-25mimic或inhibitor转染至OA软骨细胞。利用Annexin V-FITC/PI和CCK-8法检测软骨细胞细胞凋亡和细胞增殖。miR-25过表达明显抑制软骨细胞凋亡而miR-25低表达促进软骨细胞凋亡(P0.05);miR-25 mimic促进软骨细胞增殖,而miR-25 inhibitor抑制软骨细胞增殖(P0.05)。miR-25介导OA发病发生,进展中发挥着保护性调节作用,抑制OA软骨细胞凋亡和促进其增殖作用,可能对OA发病机制有新的认识和提示OA新治疗策略。     

利用miR-17的腺病毒载体抑制肺癌细胞系增殖

miR-17是肺癌的新型调节因子,有可能在肺癌发生和进展中发挥作用,但其功能尚不完全清楚。构建miR-17的腺病毒表达载体,检测miR-17对A549细胞体外增殖、锚定非依赖性生长、侵袭和转移的调控作用。利用miR-17的腺病毒载体(Ad-miR-17)在A549细胞中过表达miR-17后,使用MTT实验检测miR-17对NSCLC细胞系A549增殖作用的影响;软琼脂成集落实验检测miR-17对A549细胞锚定非依赖性生长的作用,使用Trans-well实验检测A549细胞的体外侵袭/转移作用(in vitro invasion/in vitro migration);在此基础上共转染miR-17的inhibitor确定miR-17作用的特异性。结果表明,miR-17能够显著下调A549细胞的增殖、锚定非依赖性生长、体外侵袭/转移作用;共转染miR-17的inhibitor能够阻断miR-17的作用。     

Induction of c-fos during myelomonocytic differentiation and macrophage proliferation

Previous studies have suggested a role for c-fos in cellular differentiation in fetal membranes, haematopoietic cells and teratocarcinoma stem cells. In other cell types, such as fibroblasts, c-fos expression is normally very low, but is rapidly induced by peptide growth factors, implicating c-fos in growth control mechanisms. Here, we show that the TPA (12-O-tetradecanoylphorbol-13-acetate)-induced macrophage-like differentiation of HL60 human promyelocytic precursor cells is accompanied by the induction of both c-fos mRNA and protein within 15 min after treatment, suggesting a functional role for c-fos in this differentiation system. In quiescent terminally differentiated macrophages, expression of c-fos is inducible by the macrophage-specific growth factor colony-stimulating factor-1 (CSF-1). The kinetics of c-fos induction, however, are entirely different from those in growth factor-stimulated fibroblasts, supporting the view that the c-fos gene product may serve different functions in different cell types.     

Transforming potential of the c-fms proto-oncogene (CSF-1 receptor)

The c-fms proto-oncogene encodes a transmembrane glycoprotein that is probably identical to the receptor for the macrophage colony stimulating factor, CSF-1. Forty C-terminal amino acids of the normal receptor are replaced by 11 unrelated residues in the feline v-fms oncogene product, deleting a C-terminal tyrosine residue (Tyr969) whose phosphorylation might negatively regulate the receptor kinase activity. We show that the human c-fms gene stimulates growth of mouse NIH 3T3 cells in agar in response to human recombinant CSF-1, indicating that receptor transduction is sufficient to induce a CSF-1 responsive phenotype. Although cells transfected with c-fms genes containing either Tyr969 or Phe969 were not transformed, cotransfection of these genes with CSF-1 complementary DNA induced transformation, with c-fms(Phe969) showing significantly more activity than c-fms(Tyr969). In the absence of CSF-1, chimaeric v-fms/c-fms genes encoding the wild-type c-fms C terminus were poorly transforming, whereas chimaeras bearing Phe969 were as transforming as v-fms. Thus, the Phe969 mutation, although not in itself sufficient to induce transformation, activates the oncogenic potential of c-fms in association with an endogenous ligand or in conjunction with mutations elsewhere in the c-fms gene that confer ligand-independent signals for growth.     

Expression of human proteoglycan in Chinese hamster ovary cells inhibits cell proliferation

In studying the functional role of an extracellular matrix proteoglycan, decorin, we have made observations that suggest a role for this proteoglycan in the control of cell proliferation. Extracellular matrices are made up of different combinations of collagens, elastin, hyaluronic acid, proteoglycans and various glycoproteins such as fibronectin. Most of these components can interact with cells, and much of the control of cell adhesion, migration and differentiation appears to be mediated by these interactions. Earlier studies have also attributed growth-regulatory activities to intact extracellular matrices, but the individual molecules responsible for these effects have not been characterized. We report here that Chinese hamster ovary (CHO) cell lines expressing human decorin from a stably transfected complementary DNA construct form a more orderly monolayer and grow to a lower saturation density than control cells lacking decorin. The extent of the morphological changes correlates with the level of decorin expression, and the saturation density is inversely proportional to it. The reduction in the saturation densities of the cell lines with the highest expression of decorin is more than 50%. These results reveal a novel growth inhibitory mechanism which may be related to contact inhibition of cell proliferation.     

Analysis of genes differentially expressed during initial cellular dedifferentiation in cotton

The early phase of phytohormone induction is a vital stage of somatic embryogenesis. This phase includes a key process for acquiring cellular totipotency through cellular dedifferentiation. To unravel the molecular mechanism of cellular dedifferentiation in cotton, we constructed a cDNA library using the suppression subtractive hybridization method. A total of 286 differential cDNA clones were sequenced and identified. Among these clones, 112 unique ESTs were significantly up-regulated during the early phase of phytohormone induction, and 40.2% of the ESTs were first identified. GST was highly ex- pressed from 6 to 24 h after induction with phytohormone treatment. PRPs were predominantly ex- pressed and exhibited distinct expression patterns in different treatments, suggesting that they are closely related to cellular dedifferentiation in cotton. Putative GhSAMS, GhSAMDC, GhSAHH and GhAC03 involvement in SAM metabolism was identified in this library. The analysis of qRT-PCR showed that two remarkable increased expressions of the four SAM-related genes happened during the early phase of phytohormone induction, and that a highly positive correlation existed between GhSAMS and GhSAHHo The highest expression level of GhSAMS might be associated with its reentry into the cell cycle. The histological observations further showed that some cells accomplished cellular dedifferentiation and division within 72 h in 2,4-D treatment, and that cellular dedifferentiation might be regulated through two alterations in SAM-dependent transmethylation activity in cotton. In addition, the expression patterns of differential genes in different treatments disclosed the complicated interaction between 2, 4-D and kinetin.