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1.
组织工程骨在颅颌面骨缺损临床修复中的应用   总被引:57,自引:1,他引:56  
Chai G  Zhang Y  Liu W  Cui L  Cao YL 《Zhonghua yi xue za zhi》2003,83(19):1676-1681
目的 探索人自体骨髓基质干细胞(hBMS6C)为种子细胞的组织工程骨在治疗临床颅颌面骨缺损中的可行性。方法 自1999到2002年问,选择颅颌面骨缺损病11例(外伤性颅骨缺损4例,先天性梨状孔周围骨凹陷畸形7例)进行治疗研究。从患者髂前上棘穿刺取骨髓,密度梯度离心法分离hBMSC,经体外成骨诱导和扩增至第3代。将诱导的hBMSC,与部分脱钙骨(partly demineralized bone mattix,pDBM)复合,并于体外培养一周后,手术回植骨缺损区。选择3例梨状孔凹陷畸形患者,在凹陷明显侧植入hBMSC/pDBM复合物,对侧轻度凹陷区仅植入单纯pDBM。分别于术后1,3,6,12,24,48,50个月进行临床外形和三维CT检查随访。2例患者在Ⅱ期手术时,取少量植入物活检,行组织学(HE染色),免疫组织化学(骨桥蛋白、骨粘连蛋白)检测。结果 患者三维CT检查结果示术后3~6月能形成组织工程化骨,并修复骨组织缺损。术后1~2.5年的随访表明组织工程骨稳定存在,无明显骨吸收现象,临床治疗效果稳定。组织工程骨活检标本HE染色显示其组织学结构与正常松质骨相同,并有典型软骨内化骨现象。免疫组织化学显示有骨桥蛋白、骨粘连蛋白阳性表达。而植入的单纯部分脱钙骨于术后3~6月吸收,组织学显示为脱钙骨降解碎片和纤维组织的混合物。结论 以自体hBMSC为种子细胞,利用组织工程技术可在人体内形成稳定的工程化骨组织,并临床修复颅颌面骨组织缺损。这项研究的结果为组织工程骨的临床大规模应用奠定了坚实的基础。  相似文献
2.
Background The infarct size determines the long-term prognosis of patients with acute myocardial infarction (AMI). There is a growing interest in repairing scar area by transplanting bone marrow stem cells. However, effectiveness of intracoronary injection of bone marrow mesenchymal stem cells (BMSCs) in patients with AMI still remains unclear.Methods Sixty-nine patients with AMI after percutaneous coronary intervention (PCI) were randomly divided into intracoronary injection of BMSCs (n=34) and saline (control group, n=35) groups. Serial single positron emission computer tomography (SPECT), cardiac echo and cardiac electromechanical mapping were done at the designed time intervals until six months after transplantation of BMSCs or injection of saline. Results The proportion with functional defect decreased significantly in the BMSCs patients after three months [(13±5)%] compared with that pre-transplantation [(32±11)%] and the control group [(28±10)%] at three month follow-up (P<0.05, respectively). Wall movement velocity over the infracted region increased significantly in the BMSCs group [(4.2±2.5) cm/s vs (2.2±1.3) cm/s, P<0.05], but not in the control group [(2.2±1.5) cm/s vs (2.7±1.7) cm/s, P>0.05]. Left ventricular ejection fraction (LVEF) three months after transplantation in BMSCs group increased significantly compared with that pre-implantation and with that of the control group at three months post-injection [(67±11)% vs (49±9)% and (53±8)%, P<0.05 respectively]. SPECT scan results showed that perfusion defect was improved significantly in BMSCs group at three-month follow-up compared with that in the control group [(134±66)cm2 vs (185±87)cm2, P<0.01]. At the same time, left ventricular end-diastolic volume [(136±31) ml vs (162±27) ml, P<0.05] and end-systolic volume [(63±20) ml vs (88±19) ml, P<0.05] decreased synchronously. The ratio of end-systolic pressure to end-systolic volume [Psyst/ESV, (2.84±1.30) mmHg/ml vs (1.72±1.23) mmHg/ml, P<0.05] increased significantly. Cardiac electromechnical mapping demonstrated significant improvement at three months after implantation of BMSCs compared with that pre-injection in both cardiac mechanical capability as left line local shorting [LLS, (11.29±1.64)% vs (7.32±1.86)%, P<0.05] and electrical property as left ventricular endocardial unipolar voltage [UV, (10.38±1.12) mV vs (7.61±1.09) mV, P<0.01]; perfusion defect decreased from (36.2±6.2) % to (20.3±5.31)% (P<0.01). Twenty-four-hour electrocardiographic monitoring demonstrated no arrhythmias occurred at three-months follow-up.Conclusions The transplantation of BMSCs might improve the cardiac function and it is safe and feasible with no deaths or malignant arrhythmias.  相似文献
3.
Biological features of mesenchymal stem cells from human bone marrow   总被引:55,自引:4,他引:51       下载免费PDF全文
Objective To study the biological characteristics of mesenchymal stem cells (MSCs) from human bone marrow. Methods A culture of mesenchymal stem cells was initiated from bone marrow low-density mononuclear cells separated by Percoll Centrifugation and maintained in low-glucose Dulbecco’s modified Eagle’s medium (DMEM) with 10% selected fetal calf serum. Cell growth pattern and its responses to cytokines were evaluated by trypan blue exclusion and MTT test, respectively. Cell cycle and surface antigenic features were analyzed by flow cytometry technique. Cytochemistry characteristics of MSCs were determined.Results Easy-handling methods to isolate and culture expand MSCs were developed in this study. MSCs were unique in their phenotypes. They were positive for CD29, CD44, CD166, and negative for CD34, CD45, HLA-DR and Ulex europaeus. Cytochemistry evaluation showed that MSCs were homogeneously positive for acid α-naphthl acetate esterase (ANAE), glycogen (periodic acid Schiff reaction, PAS), and negative for acid phosphatase (ACP) and the Sudan black reaction (SB). Around 5% of them were positive for alkaline phosphatase (ALP). The cells had a population doubling time of 30 hours and cell cycle analysis showed that approximately 10% of them were in S phase. MSCs grew at significantly different rates when incubated in the presence of various recombinant human cytokines, of which interferon γ, tumor necrosis factor α, stem cell factor and insulin-like growth factor promoted the proliferation of MSCs dramatically, while others tested had no effects on cell growth. Conclusions MSCs are a homogenous population of cells that have unique growth, phenotypical and cytochemical characteristics. Furthermore, the diverse responses of MSCs to different cytokines provide a clue for the selection of optimal expansion and maintenance of MSCs.  相似文献
4.
黄芩甙诱导大鼠骨髓基质细胞向神经细胞分化的研究   总被引:38,自引:1,他引:37  
Jia Y  Yang Y  Zhou Y  Song Y  Liu L  Song J  Wang X  Zhong L  Yu X 《中华医学杂志》2002,82(19):1337-1341
目的 探讨中药单体黄芩甙体外诱导大鼠骨髓基质细胞分化为神经细胞的可行性。方法 以黄芩甙作为主要诱导剂 ,诱导成年大鼠骨髓基质细胞 6d。采用免疫细胞化学染色、蛋白质印迹 (WesternBlot)、逆转录PCR(RT PCR)法检测神经细胞、胶质细胞标记蛋白和mRNA的表达 ;原位末端标记染色评价细胞凋亡率 ;绿色荧光蛋白转染后 ,进行同种异体脑移植 ,观察诱导后细胞在脑内存活情况。结果 黄芩甙诱导 6d后 ,骨髓基质细胞形成较典型的神经细胞形态。诱导前骨髓基质细胞不表达神经细胞、胶质细胞标记蛋白和mRNA ;诱导 6h ,nestin出现暂时表达 ,诱导 6d表达神经细胞标记蛋白和mRNA ,不表达胶质细胞标记蛋白和mRNA。细胞凋亡率为 12 2 %± 2 8%。同种异体脑移植 14d ,移植物存活良好。结论 黄芩甙可以定向诱导骨髓基质细胞分化为神经细胞。  相似文献
5.
目的 探讨骨髓间充质干细胞 (MSCs)局部移植对提高猪皮肤烫伤创面组织修复质量的作用 ,为临床皮肤损伤后功能性修复提供新的治疗方法。方法 抽取 6只小型香猪的骨髓 ,经体外培养并纯化MSCs,应用 5 溴脱氧尿嘧啶 (BrdU)标记技术进行标记。猪背部皮肤中线两侧各制备 6个面积为 2 5 4cm2 的深Ⅱ0 烫伤创面 ,将已标记的MSCs( 2× 10 6)以注射方式回植到提供骨髓猪的创面下 ,将创面随机分为 6组 ,即空白对照组、MSCs治疗组、MSCs 碱性成纤维细胞生长因子 (bFGF)治疗组、bFGF治疗组、MSCs 表皮细胞生长因子 (EGF)治疗组以及EGF治疗组。分别于伤后 7、14、2 1、4 2d采用大体观察、常规组织学检查、免疫组织化学及免疫荧光化学染色动态观察创面愈合情况。结果 实验猪皮肤烫伤后 7d开始 ,各组创面逐渐缩小 ,伤后 2 1d ,大部分创面愈合。虽然不同时间点各治疗组创面面积缩小率差异无显著意义 ,但以MSCs bFGF治疗组最为明显 ,伤后第 14天和 2 1天创面缩小的面积比其他 5组大 15 %~ 2 0 %。半定量评价结果显示MSCs bFGF治疗组肉芽组织中血管密度较大 ,为 ,而其他组别仅为 ~ 。再上皮化的新生表皮在MSCs bFGF治疗组较其余 5组厚 ,并有上皮角形成。半定量分析可见神经纤维在MSCs bFGF治疗组较其他 5  相似文献
6.
大鼠骨髓间充质干细胞的优化获取及生物学鉴定   总被引:23,自引:3,他引:20  
目的 优化骨髓间充质干细胞的体外获取方法并鉴定。方法 取幼年SD大鼠骨髓,分离骨髓间充质干细胞,观察细胞形态特征、生长状况及表面抗原表达。结果 分离培养的细胞有两种不同的形态,一种为小梭形细胞,一种为大扁平细胞。第9代以前生长性状稳定,增殖能力强,细胞倍增时间为48.2h;超微结构显示为早期幼稚细胞形态;免疫细胞化学检测细胞表达CD44、CD54、CD71,但不表达CD34、CD68、层粘连蛋白;在有限的传代数内,抗原表达无改变;与传统方法相比,细胞纯度高。结论 分离培养的细胞成分单一,具有干细胞特性,适用于干细胞生物学和组织工程学的研究。  相似文献
7.
组织工程技术修复犬牙槽骨缺损的实验研究   总被引:23,自引:0,他引:23  
Wang M  Weng YL  Hu XJ  Zhang Y  Chai G  Zhu L  Liu W  Cui L  Feng XP  Cao YL 《中华医学杂志》2003,83(15):1339-1344
目的 研究以犬骨髓基质细胞 (bonemarrowstromalcells ,BMSC)为种子细胞 ,利用组织工程技术修复水平型牙槽骨缺损的可行性。方法 抽取成年犬骨髓 ,用梯度离心法获取单个核细胞 ,经条件培养液体外诱导培养后 ,进行组织化学、免疫细胞化学、扫描电镜检测 ,并将培养的第 3代细胞与藻酸钙形成复合物。在犬双侧下颌第 3、4前磨牙和第 1磨牙颊侧制备冠根向深 5mm的水平型牙槽骨缺损 ,用以下方法进行治疗 :(1)细胞 藻酸钙复合物修复组 ;(2 )藻酸钙对照组 ;(3)空白对照组。4、12、2 4周后经组织学检查骨缺损修复情况。并比较 12周时实验组与对照组的修复效果。结果在体外经诱导培养的BMSC可表达AKP、cbfa1、Ⅰ型胶原等 ,表现出成骨细胞活性 ;4、12、2 4周细胞 藻酸钙复合物修复部位显示逐渐有骨形成 ;第 12周时 ,实验组牙槽嵴高度可增高 2 4mm± 0 9mm ,达到原来的 4 8 5 9% ,空白对照组增高 0 8mm± 0 8mm ,达到原来的 15 76 % ,材料对照组增高 1 0mm± 0 9mm ,达到原来的 19 74 % ,实验组的牙槽嵴增高度和修复率与两对照组均有显著性差异(P <0 0 1)。结论 能以BMSC为种子细胞 ,以藻酸钙为支架材料 ,利用组织工程技术部分修复水平型牙槽骨缺损。  相似文献
8.
小鼠c-Kit+lin-骨髓细胞移植生成肝细胞的实验研究   总被引:23,自引:2,他引:21  
目的 评价小鼠c-Kit^+Lin-骨髓细胞的肝干细胞潜能,为骨髓源性肝干细胞的临床运用提供可行性实验依据。方法 供体为纯系BALB/C雄性小鼠.c-Kit^+Lin-骨髓细胞采用免疫磁珠分选法分离细胞。受体为行35Gy全肝照射处理后的同龄同系BALB/C雌性小鼠,立即移植供体c-Kit^+lin-骨髓细胞。接受c-Kit^+lin-骨髓细胞移植1月后,活杀取肝做常规病理学检查、Y染色体的原位杂交检查、甲胎蛋白与白蛋白的免疫组化检测。结果 移植1月后小鼠肝脏的组织结构已恢复正常.肝组织细胞中存在原位杂交和免疫组化均呈阳性反应的双阳性细胞。结论 小鼠c-Kit^+Lin-骨髓细胞具有肝干细胞潜能,移植后可以在肝内定居并分化形成肝细胞。可做为肝病细胞移植疗法的种子细胞。  相似文献
9.
【目的】观察龟板含药血清体外诱导成年大鼠骨髓间充质干细胞分化为神经元的能力。【方法】采用龟板含药血清体外定向诱导第五代骨髓间充质干细胞,免疫组化鉴定神经丝蛋白(NFP)、胶质纤维酸性蛋白(GFAP)的表达。【结果】成年大鼠骨髓间充质干细胞受龟板血清诱导后神经元样细胞NF表达阳性,诱导后12h,神经元样细胞NF阳性表达达到高峰。【结论】龟板含药血清可以在体外诱导成年大鼠骨髓间充质分化为神经元。  相似文献
10.
Background In steroid-induced osteonecrosis, hypertrophy and hyperplasia of marrow fat cells and lipid deposition of osteocytes can be found in the femoral head. However, the precise reason is not clear yet. The aim of this study was to observe the effect of dexamethasone (Dex) on differentiation of marrow stromal cells (MSCs), and to investigate the pathobiological mechanism of steroid-induced osteonecrosis. Methods MSCs in cultures were treated with increasing concentrations of Dex (0, 10^-9, 10^-8, 10^-7, and 10^-6 mol/L) continuously for 21 days. The cells, which were exposed to 0 mol/L (control) or 10^-7 mol/L Dex for 4-21 days, were then cultured for 21 days without Dex. MSCs were stained with Sudan Ⅲ. Number of adipocytes was counted under a light microscope. The activity of alkaline phosphatase (ALP) of MSCs treated with 0, 10^-8, 10-7, and 10^-6 mol/L Dex for 12 days, and that treated with 0 mol/L and 10^-7 mol/L Dex for 8, 10, or 12 days were determined. The levels of triglycerides, osteocalcin and cell proliferation of MSCs treated with 0 mol/L and 10^-7 mol/L Dex were detected. The mRNA expression levels of adipose-specific 422(aP2) gene and osteogenic gene type I collagen in MSCs treated with 0 mol/L and 10^-7 mol/L Dex for 6 days were analyzed by whole-cell dot-blot hybridization. Statistical analysis was performed using Student's t test and analysis of variance. P values less than 0.05 were considered significant statistically. Results The number of adipocytes in cultures increased with the duration of MSCs' exposure to Dex and the concentration of Dex. The level of ALP activity in the MSCs decreased with concentration of Dex. In the control group, it was 8.69 times of that in the 10^-7 mol/L Dex group on day 12 (t=20.51, P〈0.001). The level of triglycerides in 10^-7 mol/L Dex group was 3.40 times of that in the control (t=11.00, P〈0.001). The levels of cell proliferation and osteocalcin in the control were 1.54 and 2.42 times of that in the 10^-7 mol/L Dex group respectively. As compared to the control, the mRNA expression of adipose-specific 422(aP2) gene in 10^-7mol/L Dex group was significantly increased (t=36.48, P〈0.001), and that of osteogenic gene type I collagen was decreased (t=42.07, P〈0.001). Conclusions Dex can directly induce the differentiation of MSCs into a large number of adipocytes and inhibit their osteogenic differentiation, which provide a novel explanation for the pathologic changes of steroid-induced osteonecrosis.  相似文献
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