苦丁茶体细胞组织培养再生植株的研究

报道了苦丁茶不经愈伤组织直接从外植体获得分化芽及愈伤组织诱导的结果，讨论了体细胞组织培养技术在苦丁茶上应用的可能性．

澳大利亚人体细胞组织治疗用品管理框架

随着人体细胞组织及其制品在临床上的日益应用,澳大利亚与其他各国一样正陆续制定相应的管理体系。本文介绍了澳大利亚人体细胞组织治疗用品的主管机构、相关的法律法规体系、人体细胞组织制品的分类及其管理等。

美国人体细胞、组织以及基于细胞和组织的产品的捐赠者资格监管研究

人体细胞、组织以及基于细胞组织的产品能够治疗许多常规药物难以治疗的疾病,这类产品正在逐渐成为热门治疗药物。由于这类产品的供体材料可能携带各种传染性病原体和疾病,因此对于供体的无风险性保障则是这类产品安全性保障的基础。关于供体安全性的保障,美国有专门的针对捐献者资格确定的法规和程序,并配套有相应的技术指南。而我国在这方面的制度设计,才刚刚开始。本文研究并借鉴美国FDA对捐献者资格确定的法规和技术指南,为我国制定保证细胞类产品供体安全性的制度提供思路和建议。

大豆体细胞胚胎发生及植株再生

以野生大豆（ＧｌｙｃｉｎｅｓｏｊａｓｉｅｂｅｔＺｕｃｃ）三个品系、半野生大豆（Ｇｌｙｃｉｎｅｇｒａｃｉｌｉｓｓｋｖ）１个品系、栽培大豆（ＧｌｙｃｉｎｅｍａｘＬＭｅｒｒｉｌ）４个品系为材料，进行组织培养．探讨了大豆体细胞组织再生植株的主要影响因素，并以器官分化和体细胞胚两种不同方式再生完整大豆植株。

苯乙酸对烟草单倍体植株叶脉组织培养一步成苗的效应研究

本研究探明了苯乙酸 (PAA)对烟草单倍体植株叶脉切段培养一步成苗的效应 ,发现 PAA与 NAA、6 - BA配合使用能有效促进一步成苗的发生。试验证明 PAA对一步成苗的促进效应与各种激素浓度配比和基本培养基的种类有关 ,与基因型的关系不明显。烟草单倍体植株叶脉切段培养一步成苗的最佳培养基配方为 MS+PAA 2 0 mg/L +NAA 0 .5 mg/L +6 - BA 1.0 mg/L +谷氨酰胺5 0 mg/L +麦芽糖 30 g/L。试验还证明 ,与 2 ,4 - D诱导产生愈伤组织途径的多级成苗培养相比 ,本研究建立的 PAA一步成苗培养具有以下优点 :1)显著提高愈伤组织的诱导率 ;2 )显著提高愈伤组织的质量 ,有效提高愈伤组织分化率 ;3)成苗率显著高于多级成苗 ;4 )再生植株具有更好的幼苗素质 ;5 )再生植株的染色体数目变异率也显著低于多级成苗 ;6 )获得双单倍体植株的频率显著高于多级成苗 ;7)能有效缩短培养周期、节约成本。

Reproduction of the Three-line Genic Male Sterile Line Parent Mian 7MB-1 （Brasscia Napus L.） and Seed Production of F1 Based on Somatic Tissue Culture

[Objective] The aim was to study the reproduction of the three-line genic male sterile （GMS） lineparent Mian7MB-1 （B. NapusL.） and the seed production of F1 through somatic tissue culture. [Methed] Through hybridization, a new breeding material Mian 7MB-1 in three-line genic temporary maintainer line propagated by tissue culture was used to improve the sterile plant rate of rapeseed in dual-purpose recessive GMS line, such as Mian 7AB type, S45AB type, and etc. And then the variety comparative test was performed. [Result] In order to avoid some fertility restoration phenomena occurring during the process of self-reproduction, Mian 7AB was propagated in bulk with somatic tissue culture of temporary maintainer line plant stem. The propagated temporary maintainer line seedlings were applied to the breeding and seed production of net room male sterile line parent, promoting the sterile plant rate of the male sterile line parent to 91.7% -93.5%. The male sterile line parents per hectare were enough for the seed production of hybrid F1 in 7 500 -15 000 hm^2. [ Conclusion ] Compared with the original dual-purpose GMS line, the seed production ultilizing male sterile line with high sterile plant rate greatly reduced the labor, significantly improved the seed yield, ensuring the seed quality and forming a perfect breeding and seed production system.

Histological Study on Soybean Somatic Embryogenesis

Soybean somatic cell could induce the development of embryoid which was similar to embryo morphologically and structurally. Somatic embryogenesis system of soybean was used to conduct genetic transformation of soybean because of its several advantages such as higher transformational efficiency, beetter synchronism and fewer plant chimeras among transgenic plants. After infected with agrobacterium tumefaciens,the initiation, differentiation and development of young cotyledon embryogenic cell of soybean which was cultured on selective culture medium with kanamycin were investigated through histological study. The result showed that somatic embryo was differentiated in non-bud differentiation way. The embryogenic cells were differentiated from epidermis of explant or cells in 1 layer or 2 layers, with the division of embryogenic cells and degradation and disorganization of surrounding cells, the embryogenic cells would form embryoid with analogous suspensor structure. Later, globular embryoid would extrude from epidermis then developed into heart-shape embryo. The experiment was expected to provide theoretical reference for the construction of high transformational system of using plant somatic embryogenesis induced by young cotyledon of soybean.

新型生物心脏瓣膜问世

新型生物心脏瓣膜问世据美国报纸报道，美国科学家经过长期努力，最近已利用人体细胞组织“培育”出心脏瓣膜，这种瓣膜移入人体后，可完全避免排异性。为了制造这种生物心脏瓣膜，科学家从患者心脏中取出一小片活体组织，并在实验室里培养，然后分离出制造瓣膜所需要的三...

生物波活性材料

生物波活性材料新颖的生物波活性材料由清华大学红典生物波工程研究中心研制成功并通过鉴定，正式投放市场。据北京市科委组织的专家鉴定，该项成果属于国内首创，达到了当前国际先进水平。上海富迪皮饰有限公司成为我国首家应用这种材料进行服装生产和销售的企业。生物波...

Nuclear β-catenin expression as a prognostic factor in advanced colorectal carcinoma

AIM： To investigate the changing pattern of β-catenin expression and its prognostic value in advanced colorectal cancer （CRC）. METHODS： Archival tumor samples were analyzed for β-catenin using immunohistochemistry （IHC） in 95 patients with advanced CRC. RESULTS： Membranous β-catenin expression was found in the normal colorectal epithelium. Almost 100% of CRC cases showed membranous and cytoplasmic expression, and 55 （58%） cases showed nuclear expression. In univariate （Kaplan-Meier） survival analysis, only the nuclear index （NI） was a significant predictor of disease free survival （DFS） （P = 0.023; n = 35）, with a NI above the median associated with longer DFS （34.2 too） than those with a NI below the median （15.5 too） （P = 0.045, ANOVA）. The other indices were not significant predictors of DFS, and none of the three tested indices （for membranous, cytoplasmic, or nuclear expression） predicted diseasespecific survival （DSS）. However, when dichotomized as positive or negative nuclear expression, the former was a significant predictor of more favorable DFS （P = 0.041） and DSS （P = 0.046）. CONCLUSION： Nuclear β-catenin expression provides additional information in predicting patient outcome in advanced CRC.

Promoting human embryonic stem cell renewal or differentiation by modulating Wnt signal and culture conditions

We previously showed that Wnt3a could stimulate human embryonic stem （hES） cell proliferation and affect cell fate determination. In the absence of feeder cell--derived factors, hES cells cultured under a feeder-free condition survived and proliferated poorly. Adding recombinant Wnt3a in the absence of feeder cell derived-factors stimulated hES cell proliferation but also differentiation. In the present study, we further extended our analysis to other Wnt ligands such as Wntl and Wnt5a. While Wntl displayed a similar effect on hES cells as Wnt3a, Wnt5a had little effect in this system. Wnt3a and Wntl enhanced proliferation of undifferentiated hES cells when feeder-derived self-renewal factors and bFGF are also present. To explore the possibility to promote the proliferation of undifferentiated hES cells by activating the Wnt signaling, we overexpressed Wnt3a or Wntl gene in immortalized human adult fibroblast （HAFi） cells that are superior in supporting long-term growth of undifferentiated hES cells than primary mouse embryonic fibroblasts. HAFi cells with or without a Wnt tmnsgene can be propagated indefinitely. Over-expression of the Wnt3a gene significantly enhanced the ability of HAFi feeder cells to support the undifferentiated growth of 3 different hES cell lines we tested. Co-expression of three commonly-used drug selection genes in Wnt3a-overpressing HAFi cells further enabled us to select rare hES clones after stable transfection or transduction. These immortalized engineered feeder cells （W3R） that co-express growth-promoting genes such as Wnt3a and three drug selection genes should empower us to efficiently make genetic modified hES cell lines for basic and translational research.

Preparation and in vitro studies of microencapsulated cells releasing human tissue inhibitor of metalloproteinase-2

Objective： To prepare microencapsulated cells releasing human tissue inhibitor ofmetalloproteinase-2 （TIMP-2）, and investigate their biological characteristics in vitro. Methods： Chinese hamster ovary （CHO） cells were stably transfected with a human TIMP-2 expression vector, encapsulated in barium alginate microcapsules and cultured in vitro. Morphological appearance of the microcapsules was observed under a light microscope. Cell viability was assessed using MTT （3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide） assay. Enzyme linked immunosorbent assay （ELISA） and reverse zymography were used to confirm the release of biologically active TIMP-2 from the microcapsules. Cryopreservation study of the microencapsulated cells was carried out using dimethyl sulfoxide （DMSO） as preservative agent. Results： The microcapsules appeared like a sphere with diameter of 300-600 ~tm. The surface of the capsule wall was clearly smooth. The microencapsulated cells survived well and kept proliferating over the 6 weeks observed. No significant difference in TIMP-2 secretion was found between encapsulated and unencapsulated cells. Reverse zymography confirmed the bioactivity of MMP （matrix metalloproteinase） inhibition of TIMP-2. The cryopreservation process did not damage the microcapsule morphology nor the viability of the cells inside. Conclusion： Microencapsulated engineered CHO cells survive at least 6 weeks after preparation in vitro, and secrete bioactive TIMP-2 freely from the microcapsules.

Aetiopathogenesis of autoimmune hepatitis

The histological hallmark of autoimmune hepatitis（AIH） is a dense portal mononuclear cell infiltrate that invades the surrounding parenchyma and comprises T and B lymphocytes,macrophages,and plasma cells.An unknown but powerful stimulus must be promoting the formation of this massive inflammatory cellular reaction that is likely to initiate and perpetuate liver damage.An autoimmune attack can follow different pathways to inflict damage on hepatocytes.Liver damage is likely to be orchestrated by CD4^＋ T lymphocytes recognizing an autoantigenic liver peptide.To trigger an autoimmune response,the peptide must be embraced by an HLA class Ⅱ molecule and presented to naive CD4^＋ T helper（Th0） cells by professional antigen presenting cells,with the co-stimulation of ligand-ligand fostering interaction between the two cells.Th0 cells become activated,differentiate into functional phenotypes according to the cytokines prevailing in the microenvironment and the nature of the antigen,and initiate a cascade of immune reactions determined by the cytokines produced by the activated T cells.Th1 cells,arising in the presence of the macrophage-derived interleukin（IL） -12,secrete mainly IL-2 and interferon-gamma（IFN-γ）,which activate macrophages,enhance expression of HLA classⅠ（increasing liver cell vulnerability to a CD8^＋ T cell cytotoxic attack）,and induce expression of HLA class Ⅱ molecules on hepatocytes.Th2 cells,which differentiate from Th0 if the microenvironment is rich in IL-4,produce mainly IL-4,IL-10,and IL-13 which favour autoantibody production by B lymphocytes.Physiologically,Th1 and Th2 antagonize each other.Th17 cells,a recently described population,arise in the presence of transforming growth factor beta（TGF-β） and IL-6 and appear to have an important effector role in inflammation and autoimmunity.Theprocess of autoantigen recognition is strictly controlled by regulatory mechanisms,such as those exerted by CD4^＋CD25^＋ regulatory T cells,which derive from Th0 in the presence of TGF-β,but in the absence of IL-6.If regulatory mechanisms fail,the autoimmune attack is perpetuated.Over the past three decades different aspects of the above pathogenic scenario have been investigated.In particular,a defect in immunoregulation affecting CD4^＋CD25^＋ regulatory T cells（T-regs） has been demonstrated in AIH,particularly at diagnosis or during relapse.Advances in the study of autoreactive T cells have occurred mostly in AIH type 2,since the knowledge that CYP2D6 is the main autoantigen has enabled the characterization of both CD4 and CD8 T cells targeting this cytochrome.CD4 T cells from patients with type 2 AIH positive for the predisposing HLA allele DRB10701 recognize seven regions of CYP2D6,five of which are also recognized by CD8 T cells.High numbers of IFN-γ producing CD4 T cells and CD8 T cells are associated with biochemical evidence of liver damage,suggesting a combined cellular immune attack.

miRNA studies in in vitro and in vivo activated hepatic stellate cells

AIM: To understand which and how different miRNAs are implicated in the process of hepatic stellate cell (HSC) activation. METHODS: We used microarrays to examine the differential expression of miRNAs during in vitro activation of primary HSCs (pHSCs). The transcriptome changes upon stable transfection of rno-miR-146a into an HSC cell line were studied using cDNA microarrays. Selected differentially regulated miRNAs were investigated by quantitative real-time polymerase chain reaction during in vivo HSC activation. The effect of miRNA mimics and inhibitor on the in vitro activation of pHSCs was also evaluated.RESULTS: We found that 16 miRNAs were upregulated and 26 were downregulated significantly in 10-d in vitro activated pHSCs in comparison to quiescent pHSCs. Overexpression of rno-miR-146a was characterized by marked upregulation of tissue inhibitor of metalloproteinase-3, which is implicated in the regulation of tumor necrosis factor-α activity. Differences in the regulation of selected miRNAs were observed comparing in vitro and in vivo HSC activation. Treatment with miR-26a and 29a mimics, and miR-214 inhibitor during in vitro activation of pHSCs induced significant downregulation of collagen type Ⅰ transcription. CONCLUSION: Our results emphasize the different regulation of miRNAs in in vitro and in vivo activated pHSCs. We also showed that miR-26a, 29a and 214 are involved in the regulation of collagen type I mRNA.

A Second Protein Marker of Caveolae:Caveolin-2

Caveolin-2,a protein about 20 kD,is a major component of the inner surface of caveolae,small invaginations of the plasma membrane.Similar with caveolin-1 and caveolin-3,it serves as a protein marker of caveolae.Caveolin-1 and-2 are located next to each other at 7q31.1 on human chromosome,the proteins encoded are co-localized and form a stable hetero-oligomeric complex,distributing similarly in tissue and cultured cells.Caveolin-3 is located on different chromosomes but confirmed to interact with caveolin-2.Caveolin-2 is similar to caveolin-1 in many respects but differs from the latter in functional domains,especially in G-protein binding domain and caveolin scaffolding domain.The mRNAs of both caveolin-1 and caveolin-2 are most abundantly expressed in white adipose tissue and are induced during differentiation of 3T3-L1 cells to adipocytes.Caveolin-2-deficinet mice demonstrate clear pulmonary defects,with little or no change in caveolin-1 expression and caveolae formation,suggesting that caveolin-2 plays a selective role in lung functions.Caveolin-2 is also involved in lipid metabolism and human cancers.

The preparation of anti-hnRNP A2/B1 polyclonal antibody and its potential application in non-small cell lung cancer

Objective： In order to evaluate potential application for diagnosis and prognosis of non-small cell-lung cancer （NSCLC）, as well as to determine its role in the pathogenesis of the disease, we prepared anti-human hnRNPA2/B1 potyclonal antibody. Methods： Prokaryotic expression vector of pET28a （＋）-hnRNP A2/B1 was constructed and bansformed into E.coli BL21. The recombinant protein induced by IPTG was purified and injected to rabbits for antibody preparation. Expression of hnRN P A2/B1 was examined in 45 tissues of NSCLC and 16 inflammatory pseudotumor tissues of lung by immunohistochemistry with the antibody. The commercial hnRNP A2/B1 monoclonal antibody was used as a controI.Results： （1） Polyclonal an-tibody against hnRNP A2/B1 with high title was obtained. （2） The positive staining in NSCLC tissues was 62.22%, which was substantially higher than that in normal tissues （40%, P = 0.035） or inflammatory pseudotumor tissues （31.25%, P=0.033）. （3） Expression of hnRNP A2/B1 positively correlated with age and the history of smoking, whereas it negatively correlated with differentiation staging of tumors. （4） Follow-up study showed that the survival time of patients with positive staining was significantly shorter than that of patients without hnRNP A2/B1 expression （P=0.048）. Conclusion： It is successful to make the recombinant protein and prepare the polyclonal antibody agonist human hnRNP A2/B1. It may be a valuable marker for the diagnosis and prognosis of NSCLC. Our results provide a basis for further study in clinical application.

Development of an Efficient Plant Regeneration System for Pelargonium×Citrosum Vanleenii

[Objective]As a mosquito-repelling ornamental plant,Pelargonium×Citrosum Vanleenii(P.× Citrosum Vanleenii)is hard to be acquired because of its hybrid background,the paper was to a new regeneration system of(P.× Citrosum Vanleenii).[Method] By studying the influence of plant growth regulators(PGRs)on explant type(leaves and petioles),the optimal combinations of PGRs to maximize SELSs(somatic embryo-like structure)and buds were established.[Result]0.2 mg/L NAA+1.0 mg/L BA was best for LS(leaves segments)and 0.2 mg/L NAA + 1.5 mg/L BAs was best for PS(petioles segments).Cultured plantlets were successfully acclimatized in soil where they grew normally without any morphological variation.Although both LS and PS were usable,the leaf was a better explant for induction of embryogenic calli,somatic embryo-like structures and buds.[Conclusion]This work offered a rapid and efficient system for plant regeneration of P.×Citrosum Vanleenii.

Pins homolog LGN regulates meiotic spindle organization in mouse oocytes

Mouse oocytes undergo polarization during meiotic maturation, and this polarization is essential for asymmetric cell divisions that maximize retention of maternal components required for early development. Without conventional centrosomes, the meiotic spindle has less focused poles and is barrel-shaped. The migration of meiotic spindles to the cortex is accompanied by a local reorganization and polarization of the cortex. LGN is a conserved protein involved in cell polarity and regulation of spindle organization. In the present study, we characterized the localization dynamics of LGN during mouse oocyte maturation and analyzed the effects of LGN upregulation and downregulation on meiotic spindle organization. At the germinal vesicle stage, LGN is distributed both cytoplasmically and at the cortex. During maturation, LGN localizes to the meiotic spindle apparatus and cortical LGN becomes less concentrated at the actin cap region. Excessive LGN induces meiotic spindle organization defects by elongating the spindle and enhancing pole focusing, whereas depletion of LGN by RNA interference results in meiotic spindle deformation and chromosome misalignment. Furthermore, the N-terminus of LGN has the ability of full-length LGN to regulate spindle organization, whereas the C-terminus of LGN controls cortical localization and polarization. Our results reveal that LGN is cortically polarized in mouse oocytes and is critical for meiotic spindle organization.