A Trabecular Bone Explant Model of Osteocyte–Osteoblast Co-Culture for Bone Mechanobiology

The osteocyte network is recognized as the major mechanical sensor in the bone remodeling process, and osteocyte-osteoblast communication acts as an important mediator in the coordination of bone formation and turnover. In this study, we developed a novel 3D trabecular bone explant co-culture model that allows live osteocytes situated in their native extracellular matrix environment to be interconnected with seeded osteoblasts on the bone surface. Using a low-level medium perfusion system, the viability of in situ osteocytes in bone explants was maintained for up to 4 weeks, and functional gap junction intercellular communication (GJIC) was successfully established between osteocytes and seeded primary osteoblasts. Using this novel co-culture model, the effects of dynamic deformational loading, GJIC, and prostaglandin E-2 (PGE(2)) release on functional bone adaptation were further investigated. The results showed that dynamical deformational loading can significantly increase the PGE(2) release by bone cells, bone formation, and the apparent elastic modulus of bone explants. However, the inhibition of gap junctions or the PGE(2) pathway dramatically attenuated the effects of mechanical loading. This 3D trabecular bone explant co-culture model has great potential to fill in the critical gap in knowledge regarding the role of osteocytes as a mechano-sensor and how osteocytes transmit signals to regulate osteoblasts function and skeletal integrity as reflected in its mechanical properties.

诱导风信子（Hyacinthus orientalis L.）同一花被外植体不同部位细胞分化花芽的研究--外源激素的作用及内源激素的变化

诱导风信子（Hyacnthus orientalis L．）同一花被外植体上不同部位细胞分化花芽，从外源激素的作用和内探激素的变化探讨细胞脱分化启动的原因，研究了不同外源激素的组合下同一花被不同部位花芽分化的诱导频率：测定了花被上、中，下三部位切块离体培养前后的内源IAA和Z+ZR的含量;在此之前研究了GC-MS．MIM内标法测定微量植物材料内源IAA含量的可行性． GC-MS．MIM内标法是测定微量（0.5-1g）植物材料内源IAA含量的一种比较理想的方法，所需材料量一般为0.58．这一方法的材料前处理采用粗提液用C18Sep-pak柱初步分离纯化．HPLC进一步纯化，能获得纯度高的样品，且操作简便，省时省力． 风信子同一花被不同部位细胞均能分化花芽．当培养基中附加2.0mg/l Zeatin或2.0mg/16-BAP时，随着外探IAA浓度从0升高到10.0mg/l．捆胞分化花芽的部位从花被下部向上部移动。 离体培养前后同一花被上、中、下三部位内源IAA和Z+ZR含量测定结果表明．风信子同一花被内源IAA含量是上部最高，下部次之，中部最低，而内源Z+ZR含量从上向下依次增加;在附加不同外源激素的MS培养基上培养3天后，同一花被上，中、下三部位内源IAA和Z+ZR含量部有一定的变化。

玉簪品种的引种，栽培和组织培养

本研究通过对玉簪属植物的组织培养，发现以腋芽、花序、试管苗的叶片为外植体，均可达到迅速繁殖的目的，但对于不同的玉簪品种来说，所能采用的外植体的种类又不尽相同：对于非嵌合体玉簪品种，以腋芽、花序、试管苗的叶片作外植体都可以;而对于嵌合体玉簪品种，如果以花序或试管苗的叶片为外植体，通过诱导不定芽进行快繁，在所得的试管苗中，超过50%的个体失去斑叶特性，这在实际生产中是不可行的，如果以腋芽为外植体，所得试管苗中多于95%的可保持原有嵌合体特征，故为理想的外植体。 在以腋芽为外植体的离体培养中，取嵌合玉簪品种H. ‘Francee’和H. ‘Ground Master’的芽作材料，通过对试验结果的分析、比较，选出了合适的繁殖培养基：MS ＋ BA0.5mg/L + NAA0.5mg/L + KH2PO4150mg/L + 水解乳蛋白500mg/L + 蔗糖30g/L + 琼脂 5.6g/L，其可提高腋芽分化率，达到了快速繁殖的目的，此外，该培养基还可保持原来的生长势，降低斑叶特性分离比;同时选出了生根培养基：MS + IAA1mg/L + 蔗糖30g/L + 琼脂5.6g/L。 以花葶为外植体的试验，证实了此种方式仅适合非嵌合体玉簪的快速繁殖。 在以叶片为外植体的研究中，取了嵌合体品种H. ‘Francee’、H. ‘Ground Master’、H. ‘Gold Standard’、H. ‘Color Glory’和H. ‘Little Ming’的再生叶征作材料，通过对试验结果的分析、比较、观察到在培养基：MS + BA4mg/L + NAA0.1mg/L + 蔗糖 30g/L + 琼脂 5.6g/L上只有芽的形成，培养基：MS + BA0.4mg/L + NAA0.4mg/L + 蔗糖 30g/L + 琼脂 5.6g/L则利于根的发生，而培养基：MS + BA4mg/L + NAA0.4mg/L + 蔗糖 30g/L + 琼脂 5.6g/L促进根或（和）不定芽的产生，则因品种的不同而变化，另外还发现，经诱导不定芽途径所得到的试管苗，仅少于5%的个体保持斑叶特性，所以不可用于嵌合体玉簪的组织培养，但可快速繁殖非嵌合体玉簪。

Linear energy transfer dependence of the effects of carbon ion beams on adventitious shoot regeneration from in vitro leaf explants of Saintpaulia ionahta

Purpose: To determine the effects of carbon ion beams with five different linear energy transfer (LET) values on adventitious shoots from in vitro leaf explants of Saintpaulia ionahta Mauve cultivar with regard to tissue increase, shoots differentiation and morphology changes in the shoots. Materials and methods: In vitro leaf explant samples were irradiated with carbon ion beams with LET values in the range of 31 similar to 151 keV/mu m or 8 MeV of X-rays (LET 0.2 keV/mu m) at different doses. Fresh weight increase, surviving fraction and percentage of the explants with regenerated malformed shoots in all the irradiated leaf explants were statistically analysed. Results: The fresh weight increase (FWI) and surviving fraction (SF) decreased dramatically with increasing LET at the same doses. In addition, malformed shoots, including curliness, carnification, nicks and chlorophyll deficiency, occurred in both carbon ion beam and X-ray irradiations. The induction frequency with the former, however, was far more than that with the X-rays. Conclusions: This work demonstrated the LET dependence of the relative biological effectiveness (RBE) of tissue culture of Saintpaulia ionahta according to 50% FWI and 50% SF. After irradiating leaf explants with 5 Gy of a 221 MeV carbon ion beam having a LET value of 96 keV/mu m throughout the sample, a chlorophyll-deficient (CD) mutant, which could transmit the character of chlorophyll deficiency to its progeny through three continuous tissue culture cycles, and plantlets with other malformations were obtained.