密骨打老儿丸含药血清对共育体系中成骨细胞和破骨细胞功能的影响

 目的： 观察密骨打老儿丸（Migu-Dalaoer pill,MDP）含药血清在成骨-破骨细胞共同培养体系中对成骨细胞（osteoblasts,OB）增殖和破骨细胞（osteoclasts,OC）骨吸收功能的影响。方法： 利用分段酶消化法从胎鼠颅骨中分离出OB,取1日龄SD大鼠四肢股骨和胫骨分离培养OC,建立培养上清相通但2种细胞间互不接触的成骨-破骨细胞共育模型。实验分为不同浓度（低、中、高）的MDP含药血清组和对照组进行比较,以细胞增殖（MTT 法）和碱性磷酸酶（alkaline phosphatase,ALP）活性代表OB的成骨活性,以抗酒石酸酸性磷酸酶（tartrate-resistant acid phosphatase,TRAP）活性和骨吸收陷窝数目代表OC的破骨能力进行测定。结果： 与对照组相比,中、高浓度MDP含药血清在成骨-破骨细胞共同培养体系中6和7 d 能显著提高OB数目和 ALP 活性（P<0.01）。与对照组相比,中、高浓度MDP含药血清在成骨-破骨细胞共同培养体系中6和7 d 能显著降低OC骨吸收陷窝的数目和分泌 TRAP 的活性（P<0.01）。结论： 密骨打老儿丸含药血清在共育体系中能够促进OB增殖和骨形成,同时抑制OC骨吸收功能。     

益骨胶囊含药血清对共育体系中破骨细胞活性和凋亡的影响

目的： 观察益骨胶囊含药血清在成骨-破骨细胞共育体系中对SD大鼠破骨细胞（OC）活性和凋亡的影响。 方法： (1)取1d龄SD大鼠颅骨分离培养成骨细胞（OB），取5d龄SD大鼠四肢股骨、胫骨分离培养OC，建立细胞上清相通但细胞间不相互混杂的平面式成骨-破骨细胞共育体系，实验分为含药血清组和对照组；（2）将10月龄SD雌性大鼠分为益骨胶囊灌胃组和生理盐水对照组，制备含药血清和对照血清；（3）重氮盐法检测抗酒石酸酸性磷酸酶（TRAP）和光镜观察骨陷窝数；（4）光镜和荧光显微镜下观察共育体系中OC凋亡情况。 结果： 含药血清组在48 h、72 h、96 h对成骨-破骨细胞共育体系中OC分泌TRACP的活性均明显降低于对照组，OC的存活数明显低于对照组，OC的凋亡率明显高于对照组且呈明显的时效关系；所形成骨吸收陷窝的数目明显低于对照组(P<0.01)。 结论： 益骨胶囊含药血清在共育体系中能够抑制OC活性，诱导破骨细胞的凋亡。     

成骨细胞参与白细胞介素—1调节破骨细胞功能的实验研究

建立了成骨细胞和破骨细胞共同培养体系，探讨了白细胞介素－１（ＩＬ１）促进骨吸收的作用机理。研究发现，破骨细胞＋ＩＬ１组的骨吸收陷窝数目和面积，与单纯破骨细胞组比较，差异无显著性意义。破骨细胞＋成骨细胞＋ＩＬ１组，骨吸收陷窝数目和面积均显著增加，与破骨细胞＋ＩＬ１组及单纯破骨细胞对照组比较，差异均有显著性意义。提示ＩＬ１对破骨细胞缺乏直接作用，而是在成骨细胞介导下，发挥调节破骨细胞的骨吸收作用。     

抗骨松丹杞颗粒含药血清对成骨-破骨细胞共培养体系中破骨细胞功能的影响

目的:观察补肾方抗骨松丹杞颗粒含药血清在成骨-破骨细胞共同培养体系中对大鼠破骨细胞(OC)骨吸收功能的影响。方法:取1 d龄SD大鼠的胎鼠颅骨与四肢骨分别分离、培养成骨细胞和破骨细胞,建立细胞上清相通但细胞间不相互混杂的平面式成骨-破骨细胞共育体系,实验分为不同浓度(低、中、高)的补肾方含药血清组和对照组进行比较,用重氮盐法检测抗酒石酸酸性磷酸酶(TRAP)和光镜观察骨陷窝数。结果:25%的补肾方含药血清组在48 h、72 h、96 h成骨-破骨细胞共育体系中OC分泌TRAP的活性明显降低于对照组;25%的补肾方含药血清组在48 h、72 h、120 h所形成骨吸收陷窝的数目明显低于对照组(P0.01)。结论:补肾方抗骨松丹杞颗粒含药血清在共育体系中能够抑制OC活性。     

阿仑膦酸钠对兔破骨细胞功能的影响

用骨陷窝形成分析法观察阿仑膦酸钠对体外培养破骨细胞功能的影响，探讨可能的作用机制。在建立兔破骨细胞培养方法的基础上，用不同浓度的阿仑膦酸钠分别与骨片或成骨细胞提前作用，然后再将骨片或成骨细胞分别与破骨细胞共同培养。结果发现当阿仑膦酸钠（10^-11,10^-9,10^-7mol/L)与骨片预处理后，破骨细胞在骨片上形成的骨吸收陷窝数目减少，其抑制率分别为19．5％（P＜0．05）、49．0％（P＜0．01）和74．5％（P＜0．01）。用阿仑膦酸钠（10^-9,10^-7,10^-5mol/L)预处理成骨细胞后，仅在高浓度（10^-5mol/L）时可见破骨细胞骨吸收陷窝的数目明显减少，其抑制率为62．8％（P＜0．01）。结果表明阿仑膦酸钠能直接或通过成骨细胞间接抑制破骨细胞的骨吸收活性。     

免疫活性细胞对骨代谢的调控作用

1 骨免疫学的概念 骨形成和骨吸收之间的平衡调节着骨内环境稳定，这包括有成骨细胞（Osteoblasts，OB）和破骨细胞（Osteoclasts，OC）间相互的协调作用。OB是骨形成细胞，可分泌骨基质分子；而来源于造血前体细胞的OC则吸收骨基质。     

益骨胶囊含药血清对共育体系中成骨细胞表达骨保护素mRNA的影响

目的 观察益骨胶囊含药血清在成骨-破骨细胞共育体系中对SD大鼠成骨细胞表达骨保护素(OPG)mRNA的影响.方法 取1d龄SD大鼠颅骨分离培养成骨细胞(OB);取5d龄SD大鼠四肢长骨分离培养破骨细胞(OC).建立上清相通但细胞间不相互混杂的平面式成骨-破骨细胞共育体系,实验分为含药血清组和对照组.将10月龄SD雌性大鼠分为益骨胶囊灌胃组和生理盐水对照组,制备含药血清和对照血清.检测共育体系中成骨细胞OPG mRNA的表达.结果 含药血清组与对照组比较,OPGmRNA表达明显升高(8.2567±0.1118比3.3350±0.9854),差异有统计学意义(P＜0.01).结论 益骨胶囊含药血清在共育体系中通过刺激OB表达OPG来实现对OC的活性和凋亡的调节.     

大鼠破骨细胞体外分离培养和鉴定

采用机械分离方法，从新生大骨长管骨分离破骨细胞获得成功。分离的破骨细胞具有如下特性：１．在体外迅速贴附于盖玻片或骨片；２．多核（一般为３￣１０个细胞核）；３．伪足运动活跃，降钙素或降钙素基因相关肽可以抑制其伪足运动；４．酸性磷酸酶染色强阳性，酸性醋酸酯酶弱阳性，ＰＡＳ中等阳性；５．细胞悬液与骨磨片共同培养，可以形成骨吸收陷窝。由此可见，本研究分离的细胞，符合破骨细胞的公认特性，故本法分离培养破骨细     

PTH对破骨细胞骨吸收功能的影响及成骨细胞介导作用

采用分离、培养兔破骨细胞和成骨细胞的方法，体外研究甲状旁腺激素（ＰＴＨ）对破骨细胞骨吸收功能的影响，以及成骨细胞和破骨细胞之间的相互作用。结果表明，ＰＴＨ对破骨细胞的骨吸收功能无直接影响，但在成骨细胞参与下，ＰＴＨ对破骨细胞性骨吸收有明显的促进作用。说明成骨细胞在ＰＴＨ调节破骨细胞功能活动中有着重要的介导作用。     

骨水泥中的硫酸钡对破骨细胞的作用

目的验证骨水泥中的硫酸钡对破骨细胞形成及其生物学活性的影响。方法体外培养外周血单核细胞并加入巨噬细胞克隆集落刺激因子及核因子B激活因子配体诱导破骨细胞分化,实验组中分别加入含有或不含有硫酸钡的骨水泥颗粒。以抗酒石酸酸性磷酸酶阳性多核细胞及象牙磨片上虫蚀样骨吸收陷窝的形成昨作为检测破骨细胞形成及其骨吸收活性的检测指标,检验骨水泥中的硫酸钡对破骨细胞的影响。结果含或不含硫酸钡的骨水泥颗粒组抗酒石酸酸性磷酸酶阳性多核细胞形成均早于无骨水泥颗粒的对照组（4天vs6天）,而骨水泥颗粒中是否含有硫酸钡的两组间抗酒石酸酸性磷酸酶阳性多核细胞形成的时间无明显差异。各组间抗酒石酸酸性磷酸酶阳性多核细胞的数量无显著差异;含硫酸钡的骨水泥颗粒组象牙磨片上骨吸收陷窝的面积较不含硫酸钡的骨水泥颗粒组及阴性对照组均增大（28.26±4.98vs22.28±3.49vs14.58±2.82,〈0.05）。结论骨水泥颗粒中的硫酸钡能够促进破骨细胞分化并促进成熟破骨细胞的骨吸收活性。     

Characterization of skin abnormalities in a mouse model of osteogenesis imperfecta using high resolution magnetic resonance imaging and Fourier transform infrared imaging spectroscopy

Evaluation of the skin phenotype in osteogenesis imperfecta (OI) typically involves biochemical measurements, such as histologic or biochemical assessment of the collagen produced from biopsy-derived dermal fibroblasts. As an alternative, the current study utilized non-invasive magnetic resonance imaging (MRI) microscopy and optical spectroscopy to define biophysical characteristics of skin in an animal model of OI. MRI of skin harvested from control, homozygous oim/oim and heterozygous oim/+ mice demonstrated several differences in anatomic and biophysical properties. Fourier transform infrared imaging spectroscopy (FT-IRIS) was used to interpret observed MRI signal characteristics in terms of chemical composition. Differences between wild-type and OI mouse skin included the appearance of a collagen-depleted lower dermal layer containing prominent hair follicles in the oim/oim mice, accounting for 55% of skin thickness in these. The MRI magnetization transfer rate was lower by 50% in this layer as compared to the upper dermis, consistent with lower collagen content. The MRI transverse relaxation time, T2, was greater by 30% in the dermis of the oim/oim mice compared to controls, consistent with a more highly hydrated collagen network. Similarly, an FT-IRIS-defined measure of collagen integrity was 30% lower in the oim/oim mice. We conclude that characterization of phenotypic differences between the skin of OI and wild-type mice by MRI and FT-IRIS is feasible, and that these techniques provide powerful complementary approaches for the analysis of the skin phenotype in animal models of disease.     

Second harmonic generation imaging microscopy studies of osteogenesis imperfecta

We have used quantitative second harmonic generation (SHG) imaging microscopy to investigate the collagen matrix organization in the oim mouse model for human osteogenesis imperfecta (OI). OI is a heritable disease in which the type I collagen fibrils are either abnormally organized or small, resulting in a clinical presentation of recurrent bone fractures and other pathologies related to collagen-comprised tissues. Exploiting the exquisite sensitivity of SHG to supramolecular assembly, we investigated whether this approach can be utilized to differentiate normal and oim tissues. By comparing SHG intensity, fibrillar morphology, polarization anisotropy, and signal directionality, we show that statistically different results are obtained for the wild type (WT) and disease states in bone, tendon, and skin. All these optical signatures are consistent with the collagen matrix in the oim tissues being more disordered, and these results are further consistent with the known weaker mechanical properties of the oim mouse. While the current work shows the ability of SHG to differentiate normal and diseased states in a mouse model, we suggest that our results provide a framework for using SHG as a clinical diagnostic tool for human OI. We further suggest that the SHG metrics described could be applied to other connective tissue disorders that are characterized by abnormal collagen assembly.     

Bone material elasticity in a murine model of osteogenesis imperfecta

To investigate the source of bone brittleness in the disease osteogenesis imperfecta (OI), biomechanical properties have been measured in the femurs from a homozygous (oim/oim) mutant mouse model of OI, its heterozygous littermates, and wild-type animals. The novel technique of ultrasound critical-angle reflectometry (UCR) was used to determine bone material elasticity matrix from measurements of the pressure and shear wave velocity at different orientations about selected points of the bone specimens. This nondestructive method is the only available means for obtaining measurements of this nature from a single surface. The ultrasound pressure wave velocity showed an increased isotropy in the homozygous compared to the wild-type specimens. This was reflected in a significant decrease in the principal elastic modulus measured along the length of the oim/oim bones (E33) while the modulus along the width (E11) did not change significantly, compared to wild-type specimens. The Poisson's ratio, v12, also had a significantly increased value in oim/oim bones. Measurements of these parameters in heterozygous animals generally fell between those from homozygous and control mice. The differences in the elasticity components in oim/oim bones indicate an altered stress distribution and a modified elastic response to loads, compared to normal bone.     

Male but not female mice with severe osteogenesis imperfecta are partially protected from high-fat diet-induced obesity

Previously we have shown that young mice with a dominant severe form of osteogenesis imperfecta (OI), caused by mutated collagen type I, exhibit an altered glucose/insulin metabolism and energy expenditure along with elevated levels of osteocalcin, a bone-derived hormone involved in the regulation of whole-body metabolism. This study aimed to examine the long-term effects of a western diet in these OI mice. Male and female OI mice and wild type littermates (WT) were fed a high-fat diet (HFD) or a matched low-fat diet (LFD) for 26 weeks. HFD-induced obesity was observed in male and female WT and female OI mice, but not in male OI mice. HFD-fed WT and OI mice of both sexes developed hyperglycemia and glucose intolerance, but the degree of glucose intolerance was significantly lower in male and female OI mice compared to sex- and diet-matched WT mice. Indirect calorimetry revealed increased movement of male OI mice on HFD compared to LFD and, while HFD lowered energy expenditure in WT mice, energy expenditure was not changed in OI mice. Further, HFD-fed male OI mice demonstrated a diet-induced increased expression of the thermogenesis genes, Ucp1 and Pgc1α, in brown adipose tissue. On LFD, total and Gla-13 osteocalcin levels were similar in 30-week-old WT and OI mice, but on HFD, both were significantly higher in OI mice than WT. Thus, male OI mice respond to HFD with increased movement, energy expenditure, brown adipose tissue thermogenesis, and higher levels of osteocalcin, resulting in partial protection against HFD-induced obesity.     

共转录因子CITED1在小鼠骨代谢中的调控作用

目的在体研究CITED1在骨代谢即成骨/破骨平衡中的调节作用,为骨质疏松的治疗提供相应的理论基础。方法利用野生型小鼠(WT小鼠)和构建的CITED1基因敲除小鼠(KO小鼠),显微电子计算机断层扫描(CT)定量测量WT小鼠和KO小鼠股骨长度、骨量和骨皮质以及骨松质厚度等骨骼表型。用ELISA检测骨代谢的血液学相关指标。用RT-qPCR检测骨标志基因的表达,从分子水平探究骨代谢改变的原因。结果 KO小鼠颅骨成骨细胞中CITED1表达极少,表明敲除成功。KO小鼠股骨长度、骨量和骨皮质以及骨松质厚度显著高于WT小鼠。KO小鼠外周血血清中I型原胶原肽(P1NP)、骨钙素(OC)和骨碱性磷酸酶(BALP)浓度均显著高于WT小鼠,而抗酒石酸酸性磷酸酶(TRAP)的浓度显著低于WT小鼠。RT-qPCR结果显示,KO小鼠颅骨成骨细胞中OC和BALP基因表达较WT小鼠显著增加(P<0. 001);同时,KO小鼠颅骨成骨细胞中抗酒石酸酸性磷酸酶(TRAP)的表达较WT小鼠显著降低(P<0. 05)。结论小鼠CITED1敲除后可以通过上调OC和BALP基因表达,下调TRAP基因的表达,促进骨形成,抑制骨吸收。     

Immature Osteoblast Lineage Cells Increase Osteoclastogenesis in Osteogenesis Imperfecta Murine

This study addressed the role of impairment of osteoblastic differentiation as a mechanism underlying pathophysiology of the osteogenesis imperfecta (OI). We hypothesized that combination of impaired osteogenic differentiation with increased bone resorption leads to diminished bone mass. By introducing visual markers of distinct stages of osteoblast differentiation, pOBCol3.6GFP (3.6GFP; preosteoblast) and pOBCol2.3GFP (2.3GFP; osteoblast/osteocytes), into the OIM model, we assessed osteoblast maturation and the mechanism of increased osteoclastogenesis. Cultures from oim/oim;2.3GFP mice showed a marked reduction of cells expressing GFP relative to +/+;2.3GFP littermates. No significant difference in expression of 3.6GFP between the +/+ and oim/oim mice was observed. Histological analysis of the oim/oim;3.6GFP mice showed an increased area of GFP-positive cells lining the endocortical surface compared with +/+;3.6GFP mice. In contrast GFP expression was similar between oim/oim;2.3GFP and +/+;2.3GFP mice. These data indicate that the osteoblastic lineage is under continuous stimulation; however, only a proportion of cells attain the mature osteoblast stage. Indeed, immature osteoblasts exhibit a stronger potential to support osteoclast formation and differentiation. We detected a higher Rankl/Opg ratio and higher expression of TNF-α in sorted immature osteoblasts. In addition, increased osteoclast formation was observed when osteoclast progenitors were cocultured with oim/oim-derived osteoblasts compared with osteoblasts derived from +/+ mice. Taken together, our data indicate that osteoblast lineage maturation is a critical aspect underlying the pathophysiology of OI.Osteogenesis imperfecta (OI) is a genetic disorder resulting in most cases from a mutation within one of the genes that encode a collagen chain. This can cause underproduction of a normal collagen molecule or secretion of a defective collagen chain that can be the basis of a disorganized and weakened matrix. Histological studies were performed to understand the bone response to the underlying genetic defect. A state of high bone turnover in the human OI was observed especially in the more severe types of OI (type III and IV).^1,2 Although the bone formation rate was higher in patients with OI, this was achieved by recruiting an increased number of osteoblasts. This histological finding would suggest that the bone formation rate per osteoblast is significantly lower in OI.¹ Thus a cellular consequence of OI can be viewed as a limitation of bone forming activity at the per cell level that is compensated by a generalized activation of the lineage to produce a greater number of osteoblastic cells. Mesenchymal stem cells that reside within the bone marrow represent a major source of osteoblasts.^3,4 Therefore, a mouse primary marrow stromal cell culture is a model most closely resembling the in vivo conditions and would allow the analysis of the osteoprogenitor cells ability to differentiate.^5,6,7To address these questions we crossed the oim/oim mouse with transgenic mice developed to identify different stages of osteoblastic differentiation. A visible marker was used to generate transgenic mice in which fragments of rat type I collagen promoter are used to drive green fluorescent protein (GFP) expression; (pOBCol3.6GFP contains 3.6-kb and pOBCol2.3GFP uses 2.3-kb region of rat Col1a1 promoter⁸). In marrow stromal cell cultures pOBCol3.6GFPtpz-positive cells first appeared at an early stage of differentiation, before nodule formation, while pOBCol2.3GFPemd fluorescence first appeared in nodules undergoing mineralization. These findings suggest that Col1a1GFP transgenes are marking different subpopulations of cells during differentiation of skeletal osteoprogenitors.^8,9 To study the biology of OI as an in vitro model that would closely resemble in vivo state we have established cultures of primary bone marrow stromal cells derived from all +/+ and oim/oim mice.In parallel to the activation of the osteoblast lineage in the oim/oim mice, the number of the osteoclasts has also been increased. In addition to increased number of osteoclasts/bone area, an increase in excretion of free DPD in urine was observed in oim/oim mice.¹⁰ A recent study by Zhang et al reported that osteoclasts in cultures derived from oim/oim mice have larger diameter, greater number of nuclei per cell, and more F-actin rings compared with +/+ osteoclasts. They also exhibited greater resorptive activity than osteoclast derived from +/+ mice.¹¹ Regulation of osteoclast precursor commitment and osteoclastic activity is indirectly regulated by the osteoblast lineage cells through the expression of the receptor activator of NF-κB ligand (Rankl) and osteoprotegerin (Opg).^12,13 In the case of OI, if much of the osteoblast lineage does not reach the stage of full differentiation this could be the underlying mechanism behind the higher induction and activity of the osteoclast lineage and the mechanism causing high bone turnover. Using previously described models, oim/oim and Col1a1GFP transgenic mice, we performed an evaluation of the in vitro and in vivo differentiation ability of osteoprogenitor cells in the oim/oim mice. We evaluated the osteoclast inductive potential of the osteoblasts derived from oim/oim mice and the ability of osteoblast lineage cells at different stages of maturation to support osteoclastogenesis. These analyses provide new insights into the mechanisms by which the genetic mutation causes a severe bone phenotype in the osteogenesis imperfecta murine (oim).     

负荷渐增式训练对老年小鼠骨骼肌卫星细胞AMPK磷酸化的影响

目的 探讨负荷渐增式训练对老年小鼠骨骼肌卫星细胞腺苷酸活化蛋白激酶（AMP-activated protein kinase，AMPK）磷酸化的影响。方法 实验小鼠分为 3 组：青年对照组（YC组，n=12）、老年对照组（OC组，n=12）与老年运动组（OT组，n=12）。OT组进行负荷渐增式训练，流式细胞分选技术分离CD45-/CD31-/Sca1-/VCAM(CD106)+细胞群体，分选细胞通过desmin、Myod肌原性染色以及成肌分化诱导培养进行肌卫星细胞鉴定，免疫组化结合Western blotting方法检测肌卫星细胞p-AMPK水平。结果 YC组骨骼肌卫星细胞AMPK及p-AMPK表达水平显著高于OC组（P<0.05）；OT组与OC组AMPK表达无明显变化（Ｐ>0.05），而OT组p-AMPK表达水平显著高于OC组（P<0.05）。结论 负荷渐增式训练可促进老年小鼠骨骼肌卫星细胞AMPK磷酸化，改善老年小鼠骨骼肌能量代谢。     

In vitro generation of osteochondral differentiation of human marrow mesenchymal stem cells in novel collagen-hydroxyapatite layered scaffolds

Integrated, layered osteochondral (OC) composite materials and/or engineered OC grafts are considered as promising strategies for the treatment of OC damage. A novel biomimetic collagen-hydroxyapatite (COL-HA) OC scaffold with different integrated layers has been generated by freeze-drying. The capacity of the upper COL layer and the lower COL/HA layer to promote the growth and differentiation of human mesenchymal stem cells (hMSCs) into chondrocytes and osteoblasts respectively was evaluated. Cell viability and proliferation on COL and COL/HA scaffolds were assessed by the MTT test. The chondrogenic differentiation of hMSCs on both scaffolds was evaluated by glucosaminoglycan (GAG) quantification, alcian blue staining, type II collagen immunocytochemistry assay and real-time polymerase chain reaction in chondrogenic medium for 21 days. Osteogenic differentiation was evaluated by alkaline phosphatase activity assay, type I collagen immunocytochemistry staining, alizarin S staining and mRNA expression of osteogenic gene for 14 days in osteogenic medium. The results indicated that hMSCs on both COL and COL/HA scaffolds were viable and able to proliferate over time. The COL layer was more efficient in inducing hMSC chondrogenic differentiation than the COL/HA layer, while the COL/HA layer possessed the superiority on promoting hMSC osteogenic induction over either COL layer or pure HA. In conclusion, the layered OC composite materials can effectively promote cartilage and bone tissue generation in vitro and are potentially usable for OC tissue engineering.     

低温保存脂肪间充质干细胞的生物学特性评价

目的 探讨低温保存对小鼠脂肪间充质干细胞（ADMSCs)体外培养的生物学特性和分化潜能的影响。方法 分离、培养小鼠ADMSCs,取第3代细胞置于液氮深低温（-196℃）冻存 12个月后复苏。MTT 法测定ADMSCs的增殖活性;β-半乳糖苷酶(SA-β-Gal)染色检测衰老;免疫细胞化学方法检测细胞表面分子CD73、CD90和CD105;条件培养基诱导后,免疫荧光法检测心肌特异性肌钙蛋白（cTnT）,茜素红、碱性磷酸酶和油红 O 染色检测成骨、成脂诱导分化潜能,Real time-PCR检测cTnT、Gata4、Ost和Runx2。未经低温保存的第3代ADMSCs为对照。 结果 低温保存的ADMSCs其增殖活性、衰老率与未冻存细胞比较差异无显著性（P>0.05）。诱导后ADMSCs的cTnT阳性表达、茜素红、碱性磷酸酶和油红O染色呈阳性反应,冻存组与对照组比较差异无显著性(P>0.05）。结论 低温保存对ADMSCs体外生长特性和成心肌、成脂肪细胞、成骨细胞的分化潜能差异无显著性。     

敲除淀粉样前体蛋白基因促进铝诱导的小鼠认知障碍损伤

目的 淀粉样前体蛋白(APP)基因是与痴呆症发生发展相关的重要基因,利用APP基因敲除小鼠探讨铝诱导的认知障碍损伤,及APP对中
毒性认知障碍损伤的作用。方法 3月龄同窝阴性小鼠分为野生对照组（WT）和铝处理组（WT+Al）,APP敲除小鼠分为模型对照组（APP^-/- ）和
模型处理组（APP^-/- +Al）,每组10只。铝处理组在粮食中加入相应剂量的乳酸铝,同窝阴性小鼠和APP^-/-小鼠给予常规鼠粮作为对照,乳酸
铝处理8周后进行水迷宫实验。HE染色观察小鼠脑组织神经病理改变;Western blotting检测糖原合成激酶3β(GSK-3β)和Caspase-3的活性变
化。结果 与WT相比,WT+Al小鼠在原平台区域停留时间和穿越原平台区域次数减少了28.1%和18.8%,而APP^-/- +Al小鼠在原平台区域停留时间
和穿越原平台区域次数减少了44.1%和51%。Western blotting显示,WT+Al小鼠和APP^-/-+Al小鼠脑组织中p-GSK-3β分别减少了17.4%和46.4%。结论 APP基因敲除促进铝诱导的神经毒性和学习记忆损伤。APP基因敲除导致GSK-3β的磷酸化水平降低、活性增高。由于GSK-3β活性增加对痴呆症具有促进作用,推测APP通过抑制GSK-3β活性在痴呆症发生过程中发挥保护效应。