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1、综上所述，我们证实淫羊藿苷刺激了人类成骨细胞的增殖和分化，这个刺激效果可能被调节， 有些部分是通过BMP2来调节的。BMPs，也就是那个属于增长因子转变因素总科的，是最初作为混合物，也就是那个引起体外或体内骨骼和软骨构造异位来鉴定的。广泛的研究已经证实bmp,包括BMP-2,是有效地区分骨组织,以及骨形成刺激物质的因素。这个家庭（BMPs2-15）中有几个成员,他们已经发现了相应的基因克隆人类互补的DNA信息库。通过重组基因技术，在充分的基础研究和临床试验中，BMPs是可供使用的。最近的研究表明rhBMP-2和rhBMP-7在各种实验系统中直接诱导了正交各向异性骨。因此,我们选择rhBMP-2
2、为淫羊藿甙的阳性对照。目前的研究表明,rhBMP-2不能刺激人的成骨细胞增殖，但可以一定浓度下通过增加ALP的活动和矿化结节的形成来诱导人类成骨细胞的分化。这一发现是与Lecanda F et al相一致的，他报告了BMP-2的存在，HMSCs的增殖和人类的造骨细胞是减少的，但是绝大多数的骨基质蛋白的mRNA含量是升高的。另外,在现在研究中最引人注目的还是本研究发现BMP-2 mRNA在成骨细胞培养的时候在淫羊藿甙的反应培养基上是增加的。造骨细胞是个骨形成细胞，是I型胶原蛋白的连续表达，ALP，骨钙素和细胞外基质的沉积钙被称为标记物的分化成骨。人类造骨细胞培养三天，在ALP的活动在1020g
3、/毫升的淫羊藿甙中表现出显著增加，在1020g /毫升的淫羊藿甙中的细胞培养了14天以后，形成的矿化结节显著增加。ALP活动的外观对于成熟的造骨细胞来说是早期表型的标志，矿化结节的形成是成骨细胞分化的后期标志，我们的结果表明，各级成骨细胞从骨先质细胞阶段到终端分化阶段，受刺激的淫羊藿甙成骨细胞是有差异的。The cell membrane surface serine/threonine kinase receptor activated on the transformation between GTP and GDP. If ligands and receptor binding too
4、 frequent, will appear two possibilities（I） receptor GTP cannot release, GDP cannot into GDP, the next activation signal cannot convey to the cells.（II）Signal system constantly release information, cells arise. Of course cells produce canceration material foundation is not only serine/threonine kina
5、se receptors. This experiment observed 20，40ng/ml barrenwort glucoside stimulation of proliferation, differentiation of ability more 10ng/ml group, possibly by the decline of causes. SMADs TGF super family downstream is signaling pathways. To the weak DPP gene alleles (BMP dominant genes) gene scree
6、ning, enhance the son found mother againstdpp (Mad) and Medea homozygous Mad variation and variation of the performance, same DPP caused the central tube, embryonic ventral and dorsal shape happen flaw. Mice and people at least 9 Mad and the homologous genes identified with Sma, and shown to form se
7、rine/threonine kinase receptor transmission path. In order to simplify the noun names, after Smad vertebrates designated as the Mad homologous gene Sma and names. SMADs in bone growth plays an important role. The common-mediator-Smad4 of SMADs recursive in very important in SAMD family members. When
8、 ligands stimulation and SMADs path restrictive phosphorylation Smad4 path with restrictive, after SMADs form hetero-oligomers. Hetero-oligomers into nuclei and activate the transcription response. In mammals, activin Smad4 and TGF- or the type of activated receptor activated Smad2 and Smad3 forming
9、 complexes, and be BMP activated receptor type of Smad1, possibly including Smad5 and Smad9 forming complexes. We concluded that the osteoblast Smad4 promote the proliferation and differentiation of role. It is worth noting,Icariine stimulate the role of osteoblast produce ALP not from Icariine oste
10、oblast contact with the then produce, but when cell proliferation, to a certain degree of in vitro osteoblast began to second differentiation Icariine stimulate the role of osteoblast produce ALP only play out. This time the role of physiological requirements with depend on.2.Extraction total RNA: A
11、bandon to morphology, join 8ml blending 5min, and add 1.6ml the chloroform：isoamyl alcohol（49:1）, violent shake, centrifugal 12000rp/m,415min；take after qing join isopropyl alcohol, centrifugal, abandon qing, precipitation，join volume fraction of 75 ethanol,10000rp/m，45min，abandon ethanol, inverted
12、centrifugal tube about 15 minutes, add 100ml DEPC treated water dissolved RNA.3. Retrovirus: take 2mg mRNA ， join 1ml Oligo（dT）18（1mg/ml）,70，immediately after 5min, add ice water bath for 5min, 5ml5xbuffer 、2.5ml dNTP （10mmol/L）、1mlRNA enzyme inhibitors（50U/ml）, 3mlAMV reverse transcriptase, 10.5mlD
13、EPC treated water, blending with 42，90min ice bath after termination of reaction.5.Reaction system, each 1ml primer,dNTP0.5ml，Taq enzymes 0.5ml, template 1ml. PCR conditions: 945min 9430s、5630s、7260s， order altogether circulation 30 times, finally 72， extensions 5min.6. Electrophoresis: 1% agarose gel electrophoresis for 40min，level plate imaging system with spectrophotometry various automatic analysis.