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By using low-frequency dynamic frequency density wave, the OA kneepad could regulate many aspects of pathological process of knee OA including: inhibition of biological effects of cytokine, MMP-3, HA, oxygen free radicals and NO; enhancing the biological activity of SOD; up-regulating the mRNA expression of Bcl-2 as well as down-regulating the mRNA expression of P53, thereby to inhibit the apoptosis of cartilage cells and delay the cartilage degeneration of the macro-morphology, cartilage cells and cartilage matrix.

OA护膝采用低频温热动态变频疏密波,针对膝OA病理过程中的多个环节进行调节,能有效抑制细胞因子、MMP-3、HA、氧自由基、NO的生物学效应,增强SOD的生物学活性,提高关节软骨细胞Bcl-2mRNA表达,减弱软骨细胞P53mRNA表达,从而抑制软骨细胞凋亡,延缓膝关节软骨宏观形态、软骨细胞及软骨基质的退变。

The blank group, the pure chondrogenic inductor group and the group of the G ranula of P enetrating Bone and Removing Pain mixed with chondrogenic inductor. We adopted pro-culture solution, pure chondrogenic induced culture solution ( TGF-β310ug/L , Dex10-7mol/L , VitC50mg/L ) and the chondrogenic induced culture solution which included the serum of the G ranula. All groups were cultivated in 50ml cell culture bottles. The effects of the G ranula of P enetrating Bone and Removing Pain on chondrogenic phenotype differentiation of BMSCs were investigated after being cultivated for 1, 2, 3 weeks, then cells observed by inverted phase contrast microscope and immunocytochemical stain.

3将第2代SD大鼠BMSCs分为空白对照组、单纯软骨诱导剂组及透骨消痛颗粒含药血清加软骨诱导剂组,采用原培养液、软骨诱导培养液(TGF-β310ug/L,Dex10-7mol/L,VitC50mg/L)及透骨消痛颗粒含药血清的软骨诱导培养液,50ml细胞培养瓶内进行培养,1、2、3周后通过倒置相差显微镜及免疫细胞化学染色等实验技术,研究透骨消痛颗粒对BMSCs诱导成软骨细胞的影响。

After 24 weeks,the operation area of group A was more smooth,and the surrounding normal cartilage naturally straight flush,transparent form new cartilage,subchondral bone formation in good condition;Group B restoration surgery the basic integrity of the cartilage tissue, center is not yet fully integrated,there was slight depression;CollagenⅡimmunohistochemistry of cartilage that was new brown area.Group C has no formation of articular cartilage.The growth and the intergration of subchondral bone of group A and B were better.

术后24周取材,见A组山羊手术区关节表面较为光滑,与周边正常软骨自然连续平齐,透明的新生软骨组织形成,软骨下骨形成完好;B组山羊手术区修复的软骨组织基本完整,中心部位仍未完全融合,有微小凹陷;Ⅱ型胶原免疫组化示新生软骨组织呈棕黄色。C组术区关节凹陷,无关节软骨组织形成。A组和B组,软骨下骨的生长及与周围组织的结合均较好,无植入物脱落现象的发生。

Coculture of autogenic BMSCs with chondrocytes can promote the proliferation of chondrocytes and the production of chondral matrix. Cocultured cells for seeding can save a number of chondrocytes, shorten culture periods and reduce subculture times.

自体BMSCs与软骨细胞共培养,BMSCs能增强软骨细胞的增殖,促进软骨细胞基质合成,缩短软骨细胞培养时间和减少传代次数,可节省大量的软骨细胞,与DBM复合后能有效修复关节软骨缺损。

The defects of articular cartilage were repaired with cartilage without chondral degenerations in group A, which was similar to the normal chondral structure.

结果:B、C组软骨损伤区形成骨纤维组织修复,术后12周,C组出现明显的软骨退行性变,B组无明显退变:A组软骨缺损区形成软骨样组织修复,且与邻近正常软骨层相连续,未见明显的软骨退变。

Staining extent and intensity were evaluated semiquantitatively and mean values for each parameter were calculated. Immunostaining with D2-40 showed positivity in 100% of skeletal myxoid chondrosarcomas, 96% of enchondromas, 95% of low-grade chondrosarcomas, 80% of chordoid meningiomas, and 75% of chordoid gliomas. Staining with S100 demonstrated diffuse, strong positivity in all (100%) chordoid gliomas, skeletal myxoid chondrosarcomas, low-grade chondrosarcomas, and enchondromas, 94% of chordomas, and 81% of extraskeletal myxoid chondrosarcomas, with focal, moderate staining in 40% of chordoid meningiomas.

我们半定量地评估了这些免疫染色的广度和强度,并且计算了它们各自的平均值。D2-40阳性表达于100%例骨的黏液样软骨肉瘤、96%例内生性软骨瘤、95%例低级别软骨肉瘤、80%例脊索样脑膜瘤和75%例脊索样胶质瘤。S100染色弥漫且强烈地表达于所有的(100%)脊索样胶质瘤、骨的黏液样软骨肉瘤、低级别软骨肉瘤和内生性软骨瘤,94%例脊索瘤,81%例骨外黏液样软骨肉瘤,还有,局灶性、中度表达于40%例黏液样脑膜瘤。

Results Among the three groups,the children's rib cartilage had the most blood vessels,the most chondrocytes,well-distributed stain of matrixes,and the type Ⅱ collagen was expressed actively and highest in photedensity.The rib cartilage of teenager group had less blood vessels,unhomogeny distributed stain of matrixes,the enlarged and separated cartilage lacunas.The rib cartilage in adult group showed the least blood vessels,the least chondrocytes.the hyalinization of perichondium,the most deposition of calcium salt,and the type II collagen was expressed at the lowest level in photodensity.

结果 儿童组肋软骨膜血管最丰富,软骨基质染色均匀,软骨细胞数目最多,Ⅱ型胶原蛋白表达最活跃,平均积分光密度值最高;青少年组软骨膜内血管减少,软骨基质染色出现明显的不均质状,软骨陷窝体积变大,并呈分隔状,陷窝内软骨细胞数目减少,II型胶原蛋白表达较儿童组减弱;成人组软骨膜血管、细胞成分明显减少,软骨膜内的纤维成分明显玻璃样变,钙盐沉积较青少年组时明显增多,Ⅱ型胶原蛋白表达较青少年组减弱。

Bone marrow-derived mesenchymal stem cells are capable of chondrogenesis, making them a possible source of cells for injectable cartilage tissue engineering. There exist different ideas on the ability of mesenchymal stem cells's chondrogenesis in monolayer culture. Because of this, the effect of adult rabbit's bone marrow-derived mesenchymal stem cells chondrogenesis in monolayer culture was studied. The mesenchymal stem cells was isolated from adult rabbit's bone marrow and monolayer cultured. TGF-β1, Vit-C and Dexamethasone were used. Immunohistochemistry analyses and histological staining of H-E, Methylaniline blue and Alcian blue were performed to identify the expression of collagen type Ⅱ and cartilage associated matrix. The results showed that the induced cells expressed and produced collagen type Ⅱ and cartilage associated matrix. This suggests that the differentiation of adult rabbit's marrow-derived mesenchymal stem cells into chondrocyte in monolayer culture is feasible and may be induced by TGF-β1, Vit-C and Dexamethasone.

骨髓基质干细胞的软骨分化潜能使其可能成为可注射组织工程化软骨研究的种子细胞,为探讨体外培养的骨髓基质干细胞在平面诱导条件下软骨分化的可行性,我们进行下列实验:获取并体外平面培养成体兔骨髓基质干细胞,应用TGF-β〓、Vit-C和地塞米松对其软骨分化诱导,诱导后的骨髓基质干细胞行细胞爬片组织学和Ⅱ型胶原免疫组化,结果证实,诱导后骨髓基质干细胞可分泌Ⅱ型胶原,组织学染色可见类似于软骨细胞,由此证明体外培养的骨髓基质干细胞在平面诱导条件下可以软骨分化,其软骨诱导因子为TGF-β〓、Vit-C和地塞米松。

The repair results of defects were observed after 4, 12 and 24 weeks. RESULTS: In experimental group, no obvious new cartilage formation was seen 4 weeks after operation; some new cartilage formation was found after 12 weeks. Histological observation showed that chondrocytes had irregular edge, honeycombing structure and that cartilage cavities formed around the chondrocytes.

结果 实验组术后 4周缺损表面未见明显的新生软骨形成;12周,缺损表面有少量的新生软骨形成,组织学切片上可见软骨细胞呈边缘不规则的,小蜂窝状结构,软骨细胞周围有软骨陷窝形成;2 4周,缺损表面的新生软骨较 4、12周的新生软骨明显,表面光滑,且与周围组织结合紧密,但仍存在部分缺损尚未修复。

Results 1、 Generally, we can see the original blue and white, shiny, no cracks in the articular surface of the cartilage after the stress increases gradually yellow, surface roughness, cracks appear; when the pressure decreases, the yellowing, rough, the color of the fracture restore gradually and become shiny.2、the shiny smooth surface can be seen under a light microscope, formation, cell distribution, tidy, clear the level of cartilage at the articular surface stress increases, the surface roughness changes, defects, disordered cells, uneven dyeing ; when the articular surface of the pressure gradually decreased, the cartilage gradually repair and the surface of cells at the surface appear only disorder.3、immunohistochemical observation can be seen throughout the observation period, cartilage cells are type Ⅱ collagen expression and expression after 3 weeks gradually weakening, when the seventh week begin to strong gradually.4、 electron microscopy shows that when stress increases the articular surface, the cartilage cells became flat, the cytoplasm in the endoplasmic reticulum, Golgi apparatus decreased with collagen disorders; and when stress decreases the articular surface, cartilage cells gradually returned normal, cytoplasm in the endoplasmic reticulum, Golgi body gradually restore quantity; collagen fibers with a gradual rules.

结果:①大体观察可见到原本蓝白色、有光泽、无裂纹的软骨在关节面压力增大后,逐渐呈灰黄色,表面粗糙,出现裂隙;当压力逐渐减小后,变黄、粗糙、有裂隙的软骨颜色逐渐恢复,变得有光泽②光镜下可见表面光滑、平整,细胞分布均匀、整齐,层次清楚的软骨在关节面压力增大后,表面变粗糙、缺损,细胞排列紊乱、染色不均;当关节面压力逐渐减小后,软骨表面逐渐修复,细胞仅在表层排列紊乱③免疫组织化学观察可见整个观察期内软骨细胞胞浆内均有Ⅱ型胶原表达,术后3周内表达逐渐变弱,从第7周时开始逐渐变强。④电镜下可见当关节面压力增大后,软骨细胞逐渐变扁,胞质中内质网膜、高尔基体减少,胶原排列紊乱;当关节面压力减小,软骨细胞形态逐渐恢复正常,胞质中内质网膜、高尔基体数量逐渐恢复;胶原纤维排列逐渐有规则。

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